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Data from the National Health and Nutrition Examination Surveys. Very large increases in blood folate levels of the U.S. population occurred between 1988-1994 and 1999-2000. Small fluctuations in blood folate levels occurred over the time period 1999-2006. The median red blood cell (RBC) folate level of the U.S. population 4 years of age and older was 266 ng/mL in 2005-2006. The median serum folate level of the U.S. population 4 years of age and older was 12.2 ng/mL in 2005-2006. In 2005-2006, the prevalence of low RBC folate (less than 140 ng/mL) among U.S. women of childbearing age (15-45 years) was 4.5%. In 2005-2006, the prevalence of low serum folate (less than 3 ng/mL) among U.S. women of childbearing age was 0.5%. Folate is an essential vitamin for good health. Women of childbearing age are among the population subgroups that have been shown previously to have low blood folate levels. Low blood folate levels are associated with an increased risk of neural tube birth defects. Beginning in 1998, the Food and Drug Administration (FDA) required the addition of folic acid (a form of folate) to all enriched breads, cereals, flours, corn meal, pasta products, rice, and other cereal grain products sold in the United States. Blood folate data from the National Health and Nutrition Examination Surveys (NHANES) have documented improvements in the folate status of the U.S. population after folate fortification was implemented. Red blood cell (RBC) folate measures long-term folate intake and low levels are associated with adverse health effects. Serum folate reflects recent folate intake and low levels are an early indicator of inadequate folate status. Pre- and postfortification blood folate levels of the U.S. population 4 years of age and older and prevalence of low blood folate among women of childbearing age (15-45 years) are reported.

The U.S. Food and Drug Administration is studying the need to monitor dietary supplements for mycotoxins such as total aflatoxins and ochratoxin A. An effective mycotoxin-monitoring program requires knowledge of the sampling and analytical variability associated with the determination of total aflatoxins (AF) and ochratoxin A (OTA) in dietary supplements. Three lots of ginger sold as a powder in capsule form and packaged in individual bottles were analyzed for both AF and OTA. The total variability associated with measuring AF and OTA in powdered ginger was partitioned into bottle-to-bottle, within bottle, and analytical variances. The variances were estimated using a nested design. For AF and OTA, the within-bottle variance associated with the 5 g laboratory sample size was the largest component of variability accounting for about 43% and 85% of the total variance, respectively; the analytical variance accounted for about 34% and 9% of the total variability, respectively; and the bottle-to-bottle variance accounted for about 23% and 7% of the total variance, respectively. When the total variance is converted into the coefficient of variation (CV or standard deviation relative to the mean concentration), the CV is lower for AF (16.9%) than OTA (24.7%).

To examine the relationship of race/ethnicity, language, communication with providers, and interpreter use with timeliness of illness and routine care for children in managed care Medicaid.Cross-sectional study using data from the Consumer Assessment of Health Plans Survey, which was administered to parents of children less than 15 years of age in Washington State managed care Medicaid in 2000.Predictors of interest were child's race/ethnicity, parent's primary language at home, difficulty communicating with the child's providers because of language, and need and use of interpreters. The primary outcomes were how often (always, usually, sometimes, or never) the child received illness and routine care as soon as wanted. Analyses used multivariate ordered logistic regression and model-based direct adjustment.A total of 5142 children were included. In adjusted analyses, children received illness and routine care significantly less often than wanted if the parent's primary language at home was not English; the parent ever had difficulty communicating with the provider; and the parent needed but did not always get an interpreter.Language barriers were associated with decreased timeliness of care for children in managed care Medicaid.

The accuracy, repeatability, and reproducibility characteristics of a method using multitoxin immunoaffinity column cleanup with liquid chromatography (LC) for determination of aflatoxins (AF; sum of aflatoxins B1, B2, G1, and G2) and ochratoxin A (OTA) in powdered ginseng and ginger have been established in a collaborative study involving 13 laboratories from 7 countries. Blind duplicate samples of blank, spiked (AF and OTA added) at levels ranging from 0.25 to 16.0 microg/kg for AF and 0.25 to 8.0 microg/kg for OTA were analyzed. A naturally contaminated powdered ginger sample was also included. Test samples were extracted with methanol and 0.5% aqueous sodium hydrogen carbonate solution (700 + 300, v/v). The extract was centrifuged, diluted with phosphate buffer (PB), filtered, and applied to an immunoaffinity column containing antibodies specific for AF and OTA. After washing the column with water, the toxins were eluted from the column with methanol, and quantified by high-performance LC with fluorescence detection. Average recoveries of AF from ginseng and ginger ranged from 70 to 87% (at spiking levels ranging from 2 to 16 microg/kg), and of OTA, from 86 to 113% (at spiking levels ranging from 1 to 8 microg/kg). Relative standard deviations for within-laboratory repeatability (RSDr) ranged from 2.6 to 8.3% for AF, and from 2.5 to 10.7% for OTA. Relative standard deviations for between-laboratory reproducibility (RSDR) ranged from 5.7 to 28.6% for AF, and from 5.5 to 10.7% for OTA. HorRat values wereor = 2 for the multi-analytes in the 2 matrixes.

Preparations of blue cohosh (Caulophyllum thalictroides) have been used traditionally by Native Americans for medicinal purposes. Dietary supplements containing dried roots or extracts of blue cohosh rhizomes are available as dietary supplements. The safety and efficacy of these preparations have not been systematically evaluated. Recent studies indicate that ingestion of specific alkaloids in blue cohosh preparations can produce birth defects and neonatal heart failure. Blue cohosh also contains saponins, which may be responsible for uterine-stimulating effects. We determined the amounts of major alkaloids and saponins in preparations of blue cohosh by high-performance liquid chromatography (HPLC). Alkaloids and saponins were monitored with a photodiode array detector and an evaporative light-scattering detector, respectively. Profiles were compared with those of authenticated blue cohosh root extracts. Identities of the alkaloids and saponins were confirmed by HPLC/mass spectrometry and nuclear magnetic resonance spectrometry. Calculations based on the results of analyses of dietary supplements showed that maximum daily intake of alkaloids and saponins will vary with the form (e.g., root, liquid extract) and doses recommended in product labeling. Intakes may vary from1 to 75 mg/day for alkaloids and from about 9 to 420 mg/day for saponins.

The effects of dietary tin on copper status and on enzymes and metabolites involved in hepatocellular antioxidant protection were measured in rats fed copper-adequate or copper-deficient diets with glucose or fructose. Rats became copper-depleted after 4 weeks on diets containing less than 0.5 micrograms of copper/g as evidenced by significant decreases in liver copper and serum ceruloplasmin. Signs of copper deficiency occurred in copper-depleted rats fed diets containing 100 micrograms of tin/g. Significant effects of tin on liver glutathione peroxidase and superoxide dismutase activities and on liver iron and total glutathione concentrations were observed. Interactions between copper and tin on liver copper and iron and on liver superoxide dismutase and malondialdehyde production are reported. Adverse effects of feeding diets containing 100 micrograms of tin/g include (i) copper depletion in rats fed copper-adequate diets, (ii) accelerated development of copper deficiency in rats fed copper-deficient diets, and (iii) reduction in hepatocellular antioxidant protection.

Conditions were optimized for the simultaneous, alkaline, aqueous methanol extraction of aflatoxins (AFL), i.e., B1 (AFB1), B2 (AFB2), G1 (AFG1), and G2 (AFG2), and ochratoxin A (OTA) with subsequent purification, isolation, and determination of the toxins in ginseng and ginger. Powdered roots were extracted with methanol-0.5% NaHCO3 solution (7 + 3). After shaking and centrifugation, the supernatant was diluted with 100 mM phosphate buffer containing 1% Tween 20 and filtered through glass microfiber filter paper. The filtrate was then passed through an immunoaffinity column, and the toxins were eluted with methanol. The AFL were separated and determined by reversed-phase liquid chromatography (RPLC) with fluorescence detection after postcolumn UV photochemical derivatization. OTA was separated and determined by RPLC with fluorescence detection. Recoveries of AFL added at 2-16 ng/g and OTA added at 1-8 ng/g to ginseng were 72-80 and 86-95%, respectively. Recoveries of AFL and OTA added to ginger were similar to those for ginseng. A total of 39 commercially available ginger products from 6 manufacturers were analyzed. Twenty-six samples were found to be contaminated with AFL at 1-31 ng/g and 29 samples, with OTA at 1-10 ng/g. Ten samples contained no AFL or OTA. Ten ginseng finished products were also analyzed; 3 contained AFL at 0.1 ng/g and 4 contained OTA at levels ranging from 0.4 to 1.8 ng/g. LC/tandem mass spectrometry with multiple-reaction monitoring of 3 collisionally induced product ions from the protonated molecular ions of OTA, AFB1, and AFG1 was used to confirm the identities of the toxins in extracts of the finished products.

In 1996, U.S. Food and Drug Administration regulations mandated the fortification of enriched cereal-grain products with folic acid, thereby emphasizing the need for validated methods for total folates in foods, particularly cereal products. The AOAC Official Methods (944.12, 960.46) currently used for the analysis of folate in foods for compliance purposes are microbiological methods. When the fortification regulations were finalized, no Official AOAC or Approved AACC methods for folate in cereal-grain products were in place. The AOAC Official Method (992.05) for folic acid in infant formula does not incorporate important improvements in the extraction procedure and was not considered suitable for the analysis of folates in foods in general. A microbiological assay protocol using a trienzyme extraction procedure was prepared and submitted for comments to 40 laboratories with recognized experience in folate analysis. On the basis of comments, the method was revised to have the conjugase (gamma-glutamyl-carboxy-peptidase) treatment follow a protease treatment, to include the use of cryoprotected inoculum, and to include the spectroscopic standardization of the standard and optional use of microtiter plates. Thirteen laboratories participated in a collaborative study of 10 required and 10 optional cereal-grain products, including flour, bread, cookies, baking mixes, and ready-to-eat breakfast cereals. The majority of the participating laboratories performed the assay by the standard test tube method; others used the microtiter plate modification for endpoint quantitation with equal success. For the required products, the relative standard deviation between laboratories (RSD(R)) ranged from 7.4 to 21.6% for 8 fortified (or enriched) products compared with expected (Horwitz equation-based) values of 11-20%. RSD(R) values were higher (22.7-52.9%) for 2 unfortified cereal-grain products. For the optional products, the RSD(R) ranged from 1.8 to 11.2% for 8 fortified products. RSD(R) values were higher (27.9-28.7%) for 2 unfortified cereal-grain products. Based on the results of the collaborative study, the microbiological assay with trienzyme extraction is recommended for adoption as Official First Action.

AOAC Official Method 999.14 is applicable for the determination of choline in milk and infant formulas. To date, its use has not been extended beyond these matrixes. We modified Official Method 999.14 and applied it to the determination of choline in a range of choline-containing dietary supplements. Dietary supplement tablets, capsules, wafers, softgels, liquid products, and drink powders were included. We found that the standard curve could be extended to cover a wider range of choline concentrations and defined a procedure for the use of Norit for samples in which the vitamin C content was high enough to interfere with the analysis. Recoveries of choline added to infant formula powders and to representative dietary supplement tablets, capsules, powdered drink mix, and wafer products were 85-114%. The use of Norit during the procedure did not affect the recovery of choline added to infant formula powders or to dietary supplements. An alkaline digestion was included for use with a product containing lecithin as the sole source of choline. Ten of 11 dietary supplement products analyzed by the modified method contained amounts of choline at or above declarations found on the product labels. The remaining product contained about 40% of the label-declared amount of choline.

Background.—Children covered by Medicaid are at increased risk of emergency department (ED) utilization. Objective.—To examine whether an association exists between ED use and perceived delay in accessing acute care. Design and Setting.—Cross-sectional study. We used data from the Consumer Assessment of Health Plans Survey collected by Washington State Medicaid in 2000. We used multivariate logistic regression to determine the odds of any ED use, using a model that included whether the parent reported not always receiving illness care for the child as soon as wanted (perceived delay), age, gender, race/ethnicity, health status, presence of a special health care need, primary language, needing an interpreter, parental education, and having a regular provider. Subjects.—Parents of a random sample of children from 9 Medicaid managed care plans were surveyed if their children were Main Outcome Measures.—Any ED use in the past 6 months. Results.—The response rate was 56.3%, yielding 5142 subjects. Of children with illnesses in the previous 6 months, 69.1% of parents reported that their child always received care as soon as they wanted; 19.9%, 8.2%, and 2.8% reported usually, sometimes, and never, respectively. Not always (vs always) receiving illness care as soon as wanted was significantly associated with increased odds of any ED use (adjusted odds ratio: 1.79; 95% confidence interval: 1.35– 2.36). Conclusion.—Children with managed care Medicaid are more likely to use the ED if parents perceive any delay in receiving illness care.

On July 11, 2003, the U.S. Food and Drug Administration published a final rule amending its food-labeling regulations to require that trans FA be declared in the nutrition label of conventional foods and dietary supplements. The effective date of this final rule is January 1, 2006. This places some urgency on increasing the number and types of currently available foods for which there are trans-fat data. Compositional databases on trans fat content of food are currently limited. The purpose of this study was to determine the trans-fat content of a wide range of foods prior to the effective date of the new regulation. AOAC Official Method of Analysis 996.01 was modified for the analysis of trans fat in noncereal products. Food products for analysis were selected on the basis of market share and data from the USDA's 1994-1996 Continuing Survey of Food Intake by Individuals. Foods were purchased from local supermarkets, weighed, hydrolyzed, converted to FAME, and analyzed by GC. The results showed that trans fat (g/100 g fat) ranged from 0.0 to 48.8 in bread, cake, and related products; from 14.9 to 27.7 in margarines; from 7.7 to 35.3 in cookies and crackers; from 24.7 to 38.2 in frozen potatoes; from 0.0 to 17.1 in salty snacks; from 0.0 to 13.2 in vegetable oils and shortenings; from 0.0 to 2.2 in salad dressings and mayonnaises; and from 0.0 to 2.0 in dry breakfast cereals. Serving sizes for the foods included in this survey ranged from 12 to 161 g, and trans-fat levels ranged from 0.0 to 7.2 g/serving. The significant differences in trans-fat content in products within each food category are due to differences in the type of fats and oils used in the manufacturing processes.

Glycoprotein IIb-IIIa is an abundant platelet receptor of the integrin family that plays a primary role in platelet aggregation. It exists on the platelet surface predominantly in a resting or inactive conformation that is converted to an active binding competent conformation upon platelet activation. There is much interest in studying the difference between active and inactive GP IIb-IIIa, developing therapeutic agents targeted towards GP IIb-IIIa and developing diagnostic assays for antibodies that recognize epitopes on GP IIb-IIIa. We present here the development of a large-scale process for purifying active GP IIb-IIIa from human platelets. The procedure results in 25mg batch sizes of high purity and activity. Additionally, the effects of detergent concentration and impurities such as IgG on ELISA assays are examined.

The cDNA that encodes the proenzyme form of human fibroblast collagenase (proMMP-1) was expressed in the methylotrophic yeast Pichia pastoris. The proMMP-1 encoding DNA was fused to the Saccharomyces cerevisiae pre-pro alpha-mating factor secretion signal in the P. pastoris pPIC9 expression plasmid, transformed into strain GS115 (His-), and His+ Muts (slow methanol utilization) transformants were selected. Full-length proenzyme and processed forms of the protein could be detected in yeast culture supernatants following shake flask and 10-liter fermentations. The protein was purified to greater than 95% homogeneity. The recombinant proMMP-1 was comparable to the native fibroblast material based on (i) migration of the full-length molecule as a 52-kDa protein on reducing SDS-PAGE, (ii) correct N-terminal amino acid sequence, (iii) activation of the full-length molecule by 4-amino-phenylmercuric acetate to yield processed protein species, (iv) degradation of gelatin as monitored by zymogram gels, and (v) enzymatic activity. These data suggest that the P. pastoris expression system offers a convenient and efficient means to produce and purify MMP-1.

A novel method for the simultaneous determination of aloin A and aloe emodin in products labeled to contain Aloe vera has been developed and validated. This analytical procedure involved extracting a test portion of the product in acetonitrile : water (40 : 60, v/v), followed by centrifugation and filtration through a 0.2 μm PTFE filter. Quantitation was achieved using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) with electrospray ionization (ESI) in the negative mode. The separation was carried out using a Waters Acquity UPLC BEH C18 column; 2.1 mm × 50 mm; 1.7 μm particle size. A five-minute gradient program was optimized at a flow rate of 0.5 mL min−1. Both water with 0.5% acetic acid and methanol with 0.5% acetic acid were used as mobile phases. The linear range was from 5.0 to 1000 ng mL−1 for aloin A and from 1.0 to 500 ng mL−1 for aloe emodin, with r ≥ 0.995 for both analytes. Since serious matrix effects (both ion suppression and ion enhancement) were eliminated by dilution for all samples, the recoveries for aloin A and aloe emodin ranged from 89–118%, and 74–108%, respectively, in the matrices tested. The method has been successfully applied to various products labeled to contain Aloe vera including liquids, capsules, soft gels and tabs. The amount of aloin A and aloe emodin in the samples ranged from none detected to 160.0 mg g−1 and from none detected to 3.8 mg g−1, respectively.