| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Articles |
a Author for correspondence. Fax 617-556-3166; e-mail pbagley{at}hnrc.tufts.edu
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Methods: Folates were heat-extracted from biological samples. A two-column HPLC system with four-channel coulometric electrochemical detection was used for analysis. An affinity column was used first to purify folates from the extract. Purified folates were eluted from the affinity column onto a phenyl analytical column, utilizing a switching valve, and folate forms were separated using an acetonitrile gradient.
Results: Folate forms differing in pteridine ring structure and number of glutamate chain residues were identified by retention time and characteristic response across the channels of the detector. Folates were quantified by comparison to an external calibration mixture. Limits of detection for pentaglutamyl folates ranged from 0.21 pmol for tetrahydrofolate to 0.41 pmol for 5-methyltetrahydrofolate. CVs (n = 5) for peaks containing 9–67 pmol of folate were 0.6–6.4% (within day) and 5.2–8.4% (between days). CVs (n = 5) for peaks containing 0.9–3.5 pmol folate were 5.7–16% (within day) and 8.4–13% (between days).
Conclusions: This automated HPLC system allows the simultaneous determination of polyglutamyl forms of folates from biological samples, including tetrahydrofolate, 5-methyltetrahydrofolate, formylated folates, and pteroylglutamate. The low detection limits allow analysis of folate form distribution in human samples such as erythrocytes and lymphocytes.
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Because the folate forms interconvert with changes in oxidation states and as one-carbon groups are added and removed, changes in the rates of folate-dependent reactions are reflected by changes in the pattern of distribution of folate coenzymes. Thus, the ability to discriminate between and to measure these folate coenzymes in tissues gives important information about in vivo folate metabolism (6).
We previously used an affinity/ion-pair HPLC method with diode array ultraviolet (UV) detection, developed in our laboratory, to measure folate distribution in tissues. With this method, the effects of folate deficiency (6), chronic choline deficiency (7), alcohol intake (8), and choline deficiency and methotrexate treatment (5) on folate form distribution in several tissues in rats have been examined. However, because the limits of detection using the diode array detector are in the range of ~1 nmol, gram quantities of tissue are required for folate analysis. This report describes an HPLC method that allows measurement of folate form distribution, in picomoles, in biological samples by use of a fully automated combination of affinity chromatography and HPLC with electrochemical detection.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
preparation of the folate affinity column
Bovine milk folate-binding protein (FBP) was isolated as described
previously (9)(10). One unit of FBP activity,
measured as described previously, was defined as the amount of FBP that
bound 1 pmol of pteroylglutamate (PteGlu) (9). The
isolated FBP was stored at 4 °C in 0.2 mol/L acetic acid at a
concentration in the range of 1 x 108 to
2.5 x 108 units/L (3.4–8.5 g protein/L)
until it was used to prepare the affinity column matrix. The first step
in preparing the column matrix was to adjust the pH of the FBP
suspension to pH 9.0 by rapidly adding 0.4 volumes of 1 mol/L potassium
tetraborate to 1 volume of the FBP suspension containing ~2.5 x
106 units of FBP activity, while stirring.
AffiPrep 10 Support (25 mL; Bio-Rad) was washed with 300 mL of ice-cold
50 mmol/L sodium bicarbonate. In this and subsequent washes, the matrix
was washed in a coarse-fritted glass filter funnel under vacuum. The
washed AffiPrep 10 support was added, while stirring, to the pH 9.0 FBP
suspension. After stirring slowly for 24 h at 4 °C, the slurry
was transferred back to the filter funnel. Unbound FBP, which was added
at an ~20-fold excess, was eluted from the FBP-AffiPrep 10 matrix
with 50 mL of 0.1 mol/L sodium bicarbonate. The FBP in this eluate
could be recovered with a folate-Sepharose column, as described
previously (9).
The FBP-AffiPrep 10 matrix remaining in the filter funnel was washed sequentially with 100 mL of 20 mmol/L trifluoroacetic acid, 200 mL of 1 mol/L potassium phosphate, pH 7, and 500 mL of water. This sequence of washes was repeated twice. The washed FBP-AffiPrep 10 matrix was stored at 4 °C in approximately four volumes of 0.3 g/L sodium azide.
To prepare the affinity column, the FBP-AffiPrep 10 slurry was transferred to a 20 x 4.6 mm HPLC column (Keystone Scientific) with a Pasteur pipette, and the column was packed under vacuum. The packing in the affinity column was replaced at least every 48 h, if the system was being run continually, or each time the system was started up.
affinity chromatography-hplc
The affinity chromatography and HPLC system consisted of a Gilson
ASPEC XL sample processor (used as an autosampler) with refrigerated
sample racks, a Rheodyne 7126 automatic switching valve and solenoid
7163 (Supelco), a Hewlett Packard 1100 quaternary pump with vacuum
degasser and Hewlett Packard 1100 control module, a Waters Model 510
pump, and an ESA CoulArray electrochemical detector equipped with a
Model 6210 four-sensor cell with CoulArray software for Windows.
The electrochemical detector was an array detector consisting of
four flow-through porous-carbon graphite coulometric electrodes each
set at incrementally higher potentials (against a palladium
reference electrode). Electroactive compounds, such as folates, begin
to oxidize at a specific electrode, depending on their individual
oxidation potential. Because coulometric electrodes operate at close to
100% efficiency, compounds that oxidize at the lower potential
electrodes are completely oxidized; therefore, the oxidized compounds
are not detected at higher potentials. A review on multielectrode array
detectors has been published by Svendsen (11).
The analysis of folate involved two automated chromatographic steps.
The first step consisted of loading the sample onto an FBP-Affiprep 10
affinity column and washing all non-folate compounds from the affinity
column. The second step consisted of eluting the purified folates from
the affinity column onto a 250 x 4.6 mm Betasil Phenyl analytical
column (Keystone) for separation and subsequent electrochemical
detection of the folate polyglutamates. To maintain a constant flow
over the electrochemical detector, a second pump was used to deliver
mobile phase to the analytical column and flow cells of the
electrochemical detector while the affinity column was being loaded and
washed. A switching valve between the affinity and analytical column
directed these two steps. A schematic of the chromatographic system is
given in Fig. 1
. In position one, the switching valve directs flow from the
Hewlett Packard quaternary pump (designated as pump 1) over the
affinity column to a waste line and flow from the Waters pump
(designated as pump 2) to the analytical column. In position two, the
switching valve directs flow from pump 1 over the affinity column to
the analytical column and flow from pump 2 to a waste line.
|
All mobile phases were made with HPLC-grade water and acetonitrile (both from Baker) and were filtered through a 0.2 µm filter before use. Mobile phases and their reservoirs were changed daily. Mobile phase A was 28 mmol/L dibasic potassium phosphate and 60 mmol/L phosphoric acid in water. Mobile phase B was 28 mmol/L dibasic potassium phosphate and 60 mmol/L phosphoric acid in 200 mL/L acetonitrile-800 mL/L water. Mobile phase C was 25 mmol/L potassium phosphate, pH 7.0, in 50 mL/L acetonitrile-950 mL/L water. Mobile phase D was water.
Pump 2 delivered mobile phase A at a rate of 0.7 mL/min continuously
throughout the run. The injector, pump 1, and the switching valve were
coordinated as described in Table 1
. Under initial conditions, the valve was in position 1
and pump 1 delivered mobile phase C at a rate of 0.35 mL/min. A sample
analysis was initiated by the ESA software signaling pump 1 to start a
run. The pump 1 control software then signaled the injector to inject
the sample. Sample (2 mL) was drawn up from the 4 °C sample rack and
loaded into the injection loop, a process that took 4 min. During this
time, the flow rate of mobile phase C was increased to 2 mL/min over
0.5 min, and the affinity column was equilibrated in 100% mobile phase
C at a flow rate of 2 mL/min. At 4 min, the flow rate was reduced to
0.35 mL/min and the sample was injected. A flow rate of 0.35 mL/min was
maintained for 6 min, during which time the sample folate bound to the
affinity column. Thereafter, the flow rate was increased to 2 mL/min,
and the affinity column was washed with 100% mobile phase C for 2.75
min and 100% mobile phase D for 5 min. The mobile phase was then
switched to 82% A-18% B, and after 1 min the flow rate was decreased
to 0.5 mL/min over 0.5 min, at which time the 82% A-18% B mobile
phase (the eluting buffer) reached the affinity column. The valve was
switched to position 2, and folates were eluted from the affinity
column to the analytical column by the acidic 82% A-18% B mobile
phase for 5.5 min. A linear gradient from 82% A-18% B mobile phase to
70% A-30% B was then run over 10 min, coinciding with a linear
increase in flow rate to 1 mL/min. A linear gradient from 70% A-30% B
to 45% A-55% B was then run over the next 17.5 min, followed by a
third linear gradient to 41% A-59% B over the next 12.5 min. At 75
min, the analytical run ended and the system returned to initial
conditions. Another sample analysis could be initiated immediately.
|
The cell potentials for the detector (vs a palladium reference electrode) were 0, 300, 500, and 600 mV for channels one to four, respectively. The detector was automatically zeroed at 16 min into the run, and the cell cleaning function at 700 mV per cell was performed for 1 min at the completion of each sample analysis.
preparation of folate calibrators
Polyglutamyl THF, 5-methyltetrahydrofolate (5-meTHF), and
10-formyltetrahydrofolate (10-foTHF) calibrators were prepared from
their respective pteroylpolyglutamate calibrators (Schircks
Laboratories), as described previously (12). The purity and
concentration of the each these calibrators was measured separately,
using ion-pair HPLC with UV diode array detection
(9)(12). A UV detector, monitoring absorbance at
280 nm, can be substituted for the diode array detector. PteGlu was
measured with either a spectrophotometer (Perkin-Elmer Lambda 7 UV/VIS
Spectrophotometer) or the ion-pair HPLC with UV detection. We used the
quantified folate calibrators to prepare an external calibration
mixture containing 5 nmol/L each of the pentaglutamyl forms of
THF, 5-meTHF, 10-foTHF, and PteGlu in 10 g/L sodium ascorbate.
This external calibration mixture was stored at -70 °C in 2.5-mL
aliquots in Vacutainer Tubes for up to 1 month.
A stock mixture of approximately equal concentrations of PteGlu containing one to seven glutamates was used to synthesize THF, 5-meTHF, and 10-foTHF calibrators with one to seven glutamates (12). Each of the resulting polyglutamyl calibration mixtures was injected onto an ion-pair HPLC with diode array detection to confirm the identity of the synthesized folates (9)(12).
microbial assay of total folate
Folates in extracts from rat and mouse brains were measured using
the HPLC method described above. Folates were also measured in an
aliquot of the same extract, using a Lactobacillus casei
microbial assay as described by Horne and Patterson (13)
with modifications by Tamura et al. (14). The extract (50
µL) was incubated for 2 h at 37 °C with chick pancreas
conjugase (10 µL), prepared as described by Mims and Laskowski
(15), to convert the folates to their monoglutamyl forms.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
. As can be seen in Fig. 2
, A–C, each form of folate,
regardless of glutamate chain length, had a characteristic response
across the four channels of the detector. We have found that, although
remaining characteristic, these responses could shift gradually over
time and that the response might shift with the replacement of the
electrode or the analytical column. The elution gradient was adjusted
to optimize peak height at the cost of separation of the mono-, di-,
and triglutamyl forms of 5,10-methenyltetrahydrofolate
(5,10-methenylTHF) because these folate forms are rarely found in
biological samples. Separation of these folate forms could be attained
by decreasing the slope of the gradient; however, there was substantial
loss of height of the later-eluting peaks.
|
The majority of the formylated forms of folate eluted as 5,10-methenylTHF, even when the injected calibrator was 10-foTHF, because of isomerization attributable to the acidic pH of the mobile phase during chromatography (16).
Individual folates in samples were identified and quantified by
comparison to folate calibrators. For an individual peak, the
combination of retention time and response pattern across the four
channels of the detector was used to identify the pteridine ring
chemistry and glutamate chain length of the folate producing the peak.
Retention times were compared to retention times of the polyglutamyl
folate calibrators, such as those shown in Fig. 2
, A–C.
Quantification of individual folates was based on comparison to the
external pentaglutamyl folate calibration mixture. This external
calibration mixture contained 10 pmol each of pentaglutamyl THF,
5-meTHF, 10-foTHF, and PteGlu per 2-mL injection volume. This
calibration mixture was run every 10–12 samples. For each folate form,
the sum of the integrated areas of all peaks in channels showing a
response was calculated and divided by 10 pmol to obtain a value of
total nanocoulombs per pmol for each individual pentaglutamyl folate.
For each individual sample folate, the sum of all peaks in all channels
with a response was divided by the value (in nanocoulombs per pmol)
obtained for the external folate calibrator with the same
pteridine ring chemistry as the sample folate, regardless of glutamate
chain length, to obtain a value of pmol of folate in the sample peak.
Pentaglutamyl forms of the folate calibrators were used because the
pentaglutamyl folate forms are typically the predominant form of folate
found in biological samples (4). Comparisons of monoglutamyl
folate calibrators and pentaglutamyl folate calibrators showed that
there was no differences in the peak areas between the mono- and
pentaglutamyl forms of the same folates (data not shown).
Depending on the size of the peaks, there could be coelution of
pentaglutamyl THF and monoglutamyl 5-meTHF, and heptaglutamyl THF and
diglutamyl 5-meTHF. Resolution of these folate forms was based on the
equations used previously (12). To resolve THF and 5-meTHF,
the following equations were used:
 |
 |
where Ax and Ay are the areas, in nanocoulombs, of the peaks in channels x and y, respectively; [T] and [M] are the sums of the areas in all channels attributable to THF and 5-meTHF, respectively, and ET and EM are the channel efficiencies for THF and 5-meTHF, respectively, for channel x or y, as indicated by the subscript. Channel efficiencies are the ratio of the area in an individual channel divided by the sum of areas of all channels. Typically, channel efficiencies were calculated from pure peaks in the same chromatogram, such as tetraglutamyl THF and pentaglutamyl 5-meTHF. The two channels with the most different efficiencies for THF and 5-meTHF were used for channels x and y.
The equations can be solved for [M] algebraically to give the
following equation:
 |
Representative chromatograms of folates extracted from rat liver
and cultured human lymphoblasts are shown in Fig. 3
.
|
linearity and detection limits
Injections of stock solutions containing a combination of 0.5–150
pmol of each of the pentaglutamyl forms of THF, 5-meTHF, 10-foTHF, and
PteGlu per injection indicated that the monitored signals were
linear for each analyte. The linear regression data are given in Fig. 4
. The slopes of the regression lines are measures for analytical
sensitivity. The upper limit of linearity was limited by the capacity
of the affinity column. At amounts >500 pmol of total folate, the
affinity column did not completely retain all forms of folates.
|
Peak heights of the pentaglutamyl calibrators were used to determine
detection limits. Because signals were seen in multiple channels for
each folate, the detection limit for each folate was determined by
comparing the signal and noise from the dominant channel only as well
as by comparing the sum of the peak height for all channels giving a
signal with the sum of the noise in all pertinent channels. Results are
given in Table 2
.
|
recovery of added calibrators
To determine the recovery of calibrators added to sample, the
extraction procedure, as described in Materials and Methods,
was performed on rat liver alone; a combination of rat liver plus
pentaglutamyl THF, 5-meTHF, and 10-foTHF; and a combination of
pentaglutamyl THF, 5-meTHF, and 10-foTHF. A 2-mL aliquot of each
extract, containing the equivalent of 10 mg of liver and/or 43 pmol of
THF, 75 pmol of 5-meTHF, and 75 pmol of 10-foTHF, was injected onto the
HPLC in triplicate. The recovery of each folate added to the rat liver
was calculated using the following formula:
 |
The recoveries for the pentaglutamyl forms of THF, 5-meTHF, and 10-foTHF (as 5,10-methenylTHF) were 91.7% ± 1.7%, 88.7% ± 2.4%, and 103.6% ± 3.8% (mean ± SE), respectively.
precision
Intra- and interassay precision was determined using rat liver
extracts. Rat liver was extracted (5 g/L), and five aliquots of the
extracted liver were run in succession to obtain intraassay precision.
To cover a range of folate concentrations, pentaglutamyl folates, the
predominant folate forms found in rat liver, were quantified, as well
as tetraglutamyl forms of folate, which typically are minor folate
forms in rat liver. The mean values and CVs for the tetra- and
pentaglutamyl forms of THF, 5-meTHF, and 5,10-methenylTHF were
determined, and the results are given in Table 3
.
|
Intraassay precision was determined in the same manner as interassay
precision, except that individual aliquots of the rat liver extract
were injected into Vacutainer Tubes and stored at -70 °C until just
prior to analysis by HPLC. Aliquots were run on separated days over a
3-week period. The mean values and CVs for the tetra- and pentaglutamyl
forms of THF, 5-meTHF, and 5,10-methenylTHF were determined, and the
results are given in Table 3
.
comparison of methods
The total folate content of extracts from 24 rat and mouse brains
was assessed by both the HPLC method and by the L. casei
microbial assay. Results of the comparison between the assays are shown
in Fig. 5
. Across the range of folate measured, the HPLC gave values that
were consistently 1.3-fold higher than the values obtained by microbial
assay, with a mean ratio of 1.30 ± 0.028 (mean ± SE)
between the HPLC and L. casei values.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
A multichannel electrochemical detector provides lower detection limits than UV detection while maintaining the advantages of diode array detection: identification of folate forms by their characteristic response across the channels, and the ability to distinguish between folates that coelute at the same retention time by use of an algorithm (12). The lower limits of detection offered by the electrochemical detection allow the analysis of folate form distribution in small specimens, such as human red blood cells and lymphocytes.
A disadvantage of this method is that the different formylated forms of folate cannot be distinguished because 10-foTHF is converted to 5,10-methenylTHF as a result of the low pH of the mobile phases (16).
There is an association between the total brain folate determined by the HPLC method presented here and the L. casei microbial method, in which the HPLC method consistently gave a value ~30% higher than the L. casei microbial method. Preliminary studies with standard reduced polyglutamyl and monoglutamyl folate, using HPLC for folate analysis, indicate that there is a loss of folate activity during the incubation with conjugase (data not shown). Conjugase treatment of folates is used before the L. casei microbial assay to cleave polyglutamyl forms of folate to monoglutamyl forms, the forms utilized by L. casei. The loss of activity during this incubation could account for the discrepancy between the two analyses.
There are several potential uses for this method. We have already demonstrated that the C677T polymorphism in the methylenetetrahydrofolate reductase gene produces an altered pattern of folate form distribution in red blood cells from individuals homozygous for the polymorphism compared with individuals with the wild-type genotype, indicating an in vivo alteration of folate metabolism (17). Measurement of folate form distribution may be an important tool in investigating the effect of nutritional, genetic, and pharmaceutical factors on folate homeostasis as well as in investigating the role of folate metabolism in vascular disease, colonic carcinoma, and neural tube defects. Because of the low detection limits of this method, folate form distribution could potentially be measured in human or animal biopsy samples, cells grown in cell culture, and in tissues isolated from animals.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
1 Nonstandard abbreviations: THF, tetrahydrofolate; UV, ultraviolet; FBP, folate-binding protein; PteGlu, pteroylglutamate; 5-meTHF, 5-methyltetrahydrofolate; 10-foTHF, 10-formyltetrahydrofolate; and 5,10-methenylTHF, 5,10-methenyltetrahydrofolate. 
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
-glutamate synthetase and organization, localization, and differential splicing of its gene. J Biol Chem 1996;271:13077-13087.
-glutamates of that organ. Biochemistry 1974;13:4957-4961.
[Medline]
[Order article via Infotrieve]
The following articles in journals at HighWire Press have cited this article:
|
|
 |
|
  A. Pribat, I. K. Blaby, A. Lara-Nunez, J. F. Gregory III, V. de Crecy-Lagard, and A. D. Hanson FolX and FolM Are Essential for Tetrahydromonapterin Synthesis in Escherichia coli and Pseudomonas aeruginosa J. Bacteriol., January 15, 2010; 192(2): 475 - 482. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Pribat, L. Jeanguenin, A. Lara-Nunez, M. J. Ziemak, J. E. Hyde, V. de Crecy-Lagard, and A. D. Hanson 6-Pyruvoyltetrahydropterin Synthase Paralogs Replace the Folate Synthesis Enzyme Dihydroneopterin Aldolase in Diverse Bacteria J. Bacteriol., July 1, 2009; 191(13): 4158 - 4165. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Nguyen, C. Tam, D. L O'Connor, B. Kapur, and G. Koren Steady state folate concentrations achieved with 5 compared with 1.1 mg folic acid supplementation among women of childbearing age Am. J. Clinical Nutrition, March 1, 2009; 89(3): 844 - 852. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. A Houghton, J. Yang, and D. L O'Connor Unmetabolized folic acid and total folate concentrations in breast milk are unaffected by low-dose folate supplements Am. J. Clinical Nutrition, January 1, 2009; 89(1): 216 - 220. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. D. Kalmbach, S. F. Choumenkovitch, A. P. Troen, P. F. Jacques, R. D'Agostino, and J. Selhub A 19-Base Pair Deletion Polymorphism in Dihydrofolate Reductase Is Associated with Increased Unmetabolized Folic Acid in Plasma and Decreased Red Blood Cell Folate J. Nutr., December 1, 2008; 138(12): 2323 - 2327. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. D Kalmbach, S. F Choumenkovitch, A. M Troen, R. D'Agostino, P. F Jacques, and J. Selhub Circulating folic acid in plasma: relation to folic acid fortification Am. J. Clinical Nutrition, September 1, 2008; 88(3): 763 - 768. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Kalmbach, S. Choumenkovitch, P. Jacques, A. Troen, and J. Selhub Unmetabolized folic acid in plasma: Effect of folic acid fortification FASEB J, April 1, 2007; 21(5): A104 - A104.
|
|
|
|
 |
|
 T. J. Vickers, G. Orsomando, R. D. de la Garza, D. A. Scott, S. O. Kang, A. D. Hanson, and S. M. Beverley Biochemical and Genetic Analysis of Methylenetetrahydrofolate Reductase in Leishmania Metabolism and Virulence J. Biol. Chem., December 15, 2006; 281(50): 38150 - 38158. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Poo-Prieto, D. B. Haytowitz, J. M. Holden, G. Rogers, S. F. Choumenkovitch, P. F. Jacques, and J. Selhub Use of the Affinity/HPLC Method for Quantitative Estimation of Folic Acid in Enriched Cereal-Grain Products J. Nutr., December 1, 2006; 136(12): 3079 - 3083. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Anguera, M. S. Field, C. Perry, H. Ghandour, E.-P. Chiang, J. Selhub, B. Shane, and P. J. Stover Regulation of Folate-mediated One-carbon Metabolism by 10-Formyltetrahydrofolate Dehydrogenase J. Biol. Chem., July 7, 2006; 281(27): 18335 - 18342. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. E Gabriel, J. W Crott, H. Ghandour, G. E Dallal, S.-W. Choi, M. K Keyes, H. Jang, Z. Liu, M. Nadeau, A. Johnston, et al. Chronic cigarette smoking is associated with diminished folate status, altered folate form distribution, and increased genetic damage in the buccal mucosa of healthy adults. Am. J. Clinical Nutrition, April 1, 2006; 83(4): 835 - 841. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. M. Troen, B. Mitchell, B. Sorensen, M. H. Wener, A. Johnston, B. Wood, J. Selhub, A. McTiernan, Y. Yasui, E. Oral, et al. Unmetabolized Folic Acid in Plasma Is Associated with Reduced Natural Killer Cell Cytotoxicity among Postmenopausal Women J. Nutr., January 1, 2006; 136(1): 189 - 194. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Orsomando, R. D. de la Garza, B. J. Green, M. Peng, P. A. Rea, T. J. Ryan, J. F. Gregory III, and A. D. Hanson Plant {gamma}-Glutamyl Hydrolases and Folate Polyglutamates: CHARACTERIZATION, COMPARTMENTATION, AND CO-OCCURRENCE IN VACUOLES J. Biol. Chem., August 12, 2005; 280(32): 28877 - 28884. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Goyer, E. Collakova, R. D. de la Garza, E. P. Quinlivan, J. Williamson, J. F. Gregory III, Y. Shachar-Hill, and A. D. Hanson 5-Formyltetrahydrofolate Is an Inhibitory but Well Tolerated Metabolite in Arabidopsis Leaves J. Biol. Chem., July 15, 2005; 280(28): 26137 - 26142. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. R. Davis, E. P. Quinlivan, K. P. Shelnutt, D. R. Maneval, H. Ghandour, A. Capdevila, B. S. Coats, C. Wagner, J. Selhub, L. B. Bailey, et al. The Methylenetetrahydrofolate Reductase 677C->T Polymorphism and Dietary Folate Restriction Affect Plasma One-Carbon Metabolites and Red Blood Cell Folate Concentrations and Distribution in Women J. Nutr., May 1, 2005; 135(5): 1040 - 1044. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. R. Davis, E. P. Quinlivan, K. P. Shelnutt, H. Ghandour, A. Capdevila, B. S. Coats, C. Wagner, B. Shane, J. Selhub, L. B. Bailey, et al. Homocysteine Synthesis Is Elevated but Total Remethylation Is Unchanged by the Methylenetetrahydrofolate Reductase 677C->T Polymorphism and by Dietary Folate Restriction in Young Women J. Nutr., May 1, 2005; 135(5): 1045 - 1050. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. P. Quinlivan, S. R. Davis, K. P. Shelnutt, G. N. Henderson, H. Ghandour, B. Shane, J. Selhub, L. B. Bailey, P. W. Stacpoole, and J. F. Gregory III Methylenetetrahydrofolate Reductase 677C->T Polymorphism and Folate Status Affect One-Carbon Incorporation into Human DNA Deoxynucleosides J. Nutr., March 1, 2005; 135(3): 389 - 396. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Ghandour, Z. Chen, J. Selhub, and R. Rozen Mice Deficient in Methylenetetrahydrofolate Reductase Exhibit Tissue-Specific Distribution of Folates J. Nutr., November 1, 2004; 134(11): 2975 - 2978. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. D. de la Garza, E. P. Quinlivan, S. M. J. Klaus, G. J. C. Basset, J. F. Gregory III, and A. D. Hanson Folate biofortification in tomatoes by engineering the pteridine branch of folate synthesis PNAS, September 21, 2004; 101(38): 13720 - 13725. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. P. Basten, M. H. Hill, S. J. Duthie, and H. J. Powers Effect of Folic Acid Supplementation on the Folate Status of Buccal Mucosa and Lymphocytes Cancer Epidemiol. Biomarkers Prev., July 1, 2004; 13(7): 1244 - 1249. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. H. Kim, J. Yang, P. B. Darling, and D. L. O'Connor A Large Pool of Available Folate Exists in the Large Intestine of Human Infants and Piglets J. Nutr., June 1, 2004; 134(6): 1389 - 1394. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S.-W. Choi, S. Friso, H. Ghandour, P. J. Bagley, J. Selhub, and J. B. Mason Vitamin B-12 Deficiency Induces Anomalies of Base Substitution and Methylation in the DNA of Rat Colonic Epithelium J. Nutr., April 1, 2004; 134(4): 750 - 755. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. M. Pfeiffer, Z. Fazili, L. McCoy, M. Zhang, and E. W. Gunter Determination of Folate Vitamers in Human Serum by Stable-Isotope-Dilution Tandem Mass Spectrometry and Comparison with Radioassay and Microbiologic Assay Clin. Chem., February 1, 2004; 50(2): 423 - 432. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Lathrop Stern, B. Shane, P. J. Bagley, M. Nadeau, V. Shih, and J. Selhub Combined Marginal Folate and Riboflavin Status Affect Homocysteine Methylation in Cultured Immortalized Lymphocytes from Persons Homozygous for the MTHFR C677T Mutation J. Nutr., September 1, 2003; 133(9): 2716 - 2720. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Anguera, J. R. Suh, H. Ghandour, I. M. Nasrallah, J. Selhub, and P. J. Stover Methenyltetrahydrofolate Synthetase Regulates Folate Turnover and Accumulation J. Biol. Chem., August 8, 2003; 278(32): 29856 - 29862. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Girelli, N. Martinelli, F. Pizzolo, S. Friso, O. Olivieri, C. Stranieri, E. Trabetti, G. Faccini, E. Tinazzi, P. F. Pignatti, et al. The Interaction between MTHFR 677 C->T Genotype and Folate Status Is a Determinant of Coronary Atherosclerosis Risk J. Nutr., May 1, 2003; 133(5): 1281 - 1285. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S.-W. Choi, S. Friso, G. G. Dolnikowski, P. J. Bagley, A. N. Edmondson, D. E. Smith, and J. B. Mason Biochemical and Molecular Aberrations in the Rat Colon Due to Folate Depletion Are Age-Specific J. Nutr., April 1, 2003; 133(4): 1206 - 1212. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. B. Mason Biomarkers of Nutrient Exposure and Status in One-Carbon (Methyl) Metabolism J. Nutr., March 1, 2003; 133(3): 941S - 947. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Jones and P. F. Nixon Tetrahydrofolates Are Greatly Stabilized by Binding to Bovine Milk Folate-Binding Protein J. Nutr., September 1, 2002; 132(9): 2690 - 2694. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Friso, S.-W. Choi, D. Girelli, J. B. Mason, G. G. Dolnikowski, P. J. Bagley, O. Olivieri, P. F. Jacques, I. H. Rosenberg, R. Corrocher, et al. A common mutation in the 5,10-methylenetetrahydrofolate reductase gene affects genomic DNA methylation through an interaction with folate status PNAS, April 16, 2002; 99(8): 5606 - 5611. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Friso, S.-W. Choi, D. Girelli, J. B. Mason, G. G. Dolnikowski, P. J. Bagley, O. Olivieri, P. F. Jacques, I. H. Rosenberg, R. Corrocher, et al. A common mutation in the 5,10-methylenetetrahydrofolate reductase gene affects genomic DNA methylation through an interaction with folate status PNAS, April 16, 2002; 99(8): 5606 - 5611. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
