Quantitative liquid chromatographic method using fluorescence detection for determining zearalenone and its metabolites in blood plasma and urine

The liquid chromatographic (LC) method described, suitable for use with both blood plasma and urine, is applicable for determination of zearalenone and alpha-zearalenol at levels as low as 0.5 ng/mL plasma and 5 ng/mL urine. The sample is incubated overnight with beta-glucuronidase to analyze for both conjugated and unconjugated forms of zearalenone. The next day, the sample is acidified with H3PO4, extracted with chloroform, and evaporated to dryness. The residue is dissolved in toluene and loaded onto a silica gel cartridge which is washed with toluene and eluted with toluene-acetone (88 + 12). The eluate is evaporated, and the residue is dissolved in chloroform, extracted with 0.18M NaOH, neutralized with H3PO4, and re-extracted with chloroform. The chloroform extract is evaporated, dissolved in mobile phase for LC, and injected onto a normal phase column under the following chromatographic conditions: mobile phase of water-saturated dichloromethane containing 2% 1-propanol, and fluorescence detector, excitation wave-length 236 nm, and 418 nm cut-off emission filter. Recoveries of zearalenone and its metabolites from blood plasma and urine are 80-89% in the range 2.0-10 ng standard/mL plasma, and 81-90% in the range 10-30 ng standard/mL urine. This method was used to analyze blood and urine samples from a pig fed zearalenone-contaminated feed (5 mg/kg), corresponding to 80 micrograms/kg body weight. Zearalenone was rapidly metabolized to alpha-zearalenol, which appeared in the blood only 30 min after feeding. Almost all zearalenone and alpha-zearalenol was found conjugated with glucuronic acid in both blood plasma and urine.     

Development of an improved liquid phase microextraction technique and its application in the analysis of flumetsulam and its two analogous herbicides in soil

An improved liquid phase microextraction (LPME) technique has been developed. As part of this technique, analytes were extracted into an extractant microdrop which was laid on the cone-shaped bottom of a PCR tube (polychloroprene rubber tube) but not at the needle tip of a microsyringe, and the sample vial and PCR tube were horizontally placed so that the extractant was not affected by the force of vertical orientation (gravity and floating force). The stability of the extractant microdrop increased greatly, and the selection of extractant was extended. In this work, flumetsulam and its two analogous herbicides were chosen as model analytes in investigating the feasibility of the new pretreatment method by coupling it to high-performance liquid chromatography (HPLC). Under the optimized experimental conditions, the linear range and the limits of detection (S/N = 3) were 0.01-5 microg/mL (r = 0.9997) and 0.8 ng/mL for flumetsulam, 0.002-5 microg/mL (r = 0.9994) and 0.5 ng/mL for analogue 1, and 0.002-1 microg/mL (r = 0.9993) and 0.5 ng/mL for analog 2, respectively. The inter- and intraday reproducibilities (RSD) were below 5.3 and 4.5%, respectively. Good recoveries that ranged from 79.4 to 115.0% were obtained in the analysis of real soil samples. The extraction efficiency of the improved method was 4-8 times higher than that of the conventional liquid phase microextraction method. The novel, simple, rapid, sensitive technique is very suitable for extraction of apolar and medium polar analyte in complex environmental samples.     

Residue depletion of ractopamine and its metabolites in swine tissues, urine, and serum

Ractopamine hydrochloride is a beta-adrenergic leanness-enhancing agent approved for use in swine in the United States. Depletion of ractopamine and its metabolites from animal tissues, urine, and serum is of interest for the detection of illegal use. The objectives of this study were to measure the residues of ractopamine in swine incurred samples after treatment with dietary ractopamine for 28 consecutive days. An efficient and sensitive analytical method was developed for the detection of parent ractopamine and its metabolites in swine tissues, urine, and serum by HPLC-FLD. After extraction, enzymatic digestion, and solid-phase cleanup of the samples, ractopamine residues were determined by liquid chromatography (LC) with fluorescence detector. The limits of detection (LOD) for tissues, urine, and serum were 1 ng g(-1), 0.5 ng mL(-1), and 0.5 ng mL(-1), respectively. Recoveries ranged from 70.5 to 94.5% for samples fortified at 1-50 ng g(-1) or ng mL(-1). Sixty pigs were fed twice daily for 28 consecutive days with feeds containing 18 mg kg(-1) ractopamine HCl. The residue concentrations in urine, liver, and kidney were 650.06 ng mL(-1), 46.09 ng g(-1), and 169.27 ng g(-1), respectively, compared with those in muscle, fat, and serum (4.94 ng g(-1), 3.28 ng g(-1), and 7.48 ng mL(-1), respectively) at the feeding period of 7 days. The residue concentrations at withdrawal period of 0 days in all edible tissues were lower than tolerance values established by the FDA and MRL values listed by the JECFA. These data support the withdrawal time of 0 days established by the FDA for ractopamine used as feed additive in swine.     

Enzyme immunoassay for tenuazonic acid in apple and tomato products

The Alternaria mycotoxin tenuazonic acid was derivatized with succinic anhydride and conjugated to keyhole limpet hemocyanin (KLH) and to horseradish peroxidase (HRP), respectively. The KLH conjugate was used to produce polyclonal antibodies in rabbits. A competitive direct enzyme immunoassay (EIA) for tenuazonic acid was established, which was moderately sensitive for tenuazonic acid [50% inhibition concentration (IC(50)): 320 ± 130 ng/mL] but strongly reacted with tenuazonic acid acetate (IC(50): 23.3 ± 7.5 ng/mL). Therefore, an optimized EIA protocol was established, which employed acetylation of standard and sample extract solutions. The mean standard curve detection limit (IC(30)) for tenuazonic acid acetate was 5.4 ± 2.0 ng/mL, enabling detection limits for tenuazonic acid in apple and tomato products of 25-50 ng/g (150 ng/g in tomato paste). Recoveries in a concentration range of 50-2000 ng/g were 60-130% in apple juice and tomato juice and 40-150% in other tomato products. Tenuazonic acid was detected in apple juice and tomato products from German retail shops at levels of 50-200 ng/g. In conclusion, this novel EIA for tenuazonic acid could be useful within a screening program for Alternaria mycotoxins in food.     

The fate of trans-caftaric acid administered into the rat stomach

trans-Caftaric acid is the most abundant nonflavonoid phenolic compound in grapes and wines. It occurs in chicory and is one of the bioactive components of Echinacea purpurea. In order to fill the gap of knowledge about its bioavailability in mammals, we investigated its absorption, tissue distribution, and metabolism in rats. Assuming that the stomach is a relevant site of absorption of dietary polyphenols, a solution of trans-caftaric acid was maintained in the ligated stomach of anaesthetized rats for 20 min. Intact trans-caftaric acid was detected in rat plasma at both 10 and 20 min (293 +/- 45 and 334 +/- 49 ng/mL, respectively), along with its O-methylated derivative trans-fertaric acid, whose concentration rose over time (from 92 +/- 12 to 185 +/- 24 ng/mL). At 20 min, both trans-caftaric acid and trans-fertaric acid were detected in the kidney (443 +/- 78 and 2506 +/- 514 ng/g, respectively) but not in the liver. Only trans-fertaric acid was found in the urine (33.3 +/- 12.8 microg/mL). In some rats, trans-caftaric acid was detected in the brain (180 +/- 20 ng/g).     

Development of ELISA and immunochromatographic assay for the detection of gentamicin

Competitive direct enzyme-linked immunosorbent assay (ELISA) and the immunochromatographic assay were developed using a monoclonal antibody to detect gentamicin in the animal plasma and milk. No cross-reactivity of the antibody was observed with other aminoglycosides based on competitive direct ELISA, indicating that the antibody is highly specific for gentamicin. On the basis of the standard curves, the detection limits were determined to be 0.9 ng/mL in phosphate-buffered saline (PBS), 1.0 ng/mL in plasma, and 0.5 ng/mL in milk, respectively. Recoveries of gentamicin from spiked plasma and milk at levels of 25-100 ng/mL ranged from 85 to 112%. The concentration of intramuscularly injected gentamicin was successfully monitored in the rabbit plasma through competitive direct ELISA. The detection limits were estimated to be about 6 ng/mL of gentamicin in PBS, plasma, and milk using the colloidal gold-based immunochromatographic assay, which is suitable for the simple screening of gentamicin residues in the veterinary field. Observed positives can be confirmed using a more sensitive laboratory method such as competitive direct ELISA. Therefore, the assays developed in this study could complement each other as well as veterinary field and laboratory findings.     

Online YPA4 resin microcolumn separation/preconcentration coupled with inductively coupled plasma optical emission spectrometry (ICP-OES) for the speciation analysis of mercury in seafood

A simple and effective method for the determination of trace amounts of methylmercury (MeHg(+)) and inorganic mercury (Hg(2+)) in seafood was developed by online microcolumn separation/preconcentration combined with inductively coupled plasma optical emission spectrometry (ICP-OES). It was found that Hg(2+) could be quantitatively adsorbed by YPA 4 resin from pH 7.0 to strong acidic medium (6 mol L(-1) HCl) and that MeHg(+) was retained by the YPA 4 microcolumn only at pH 1.0-7.0. Therefore, a strong acidic medium (about 5 mol L(-1) HCl), which could liberate mercury species from biological samples, was used to directly separate inorganic Hg(2+) from total Hg, and MeHg(+) in effluent was retained by YPA 4 column after the effluent was adjusted to pH 1.5. The effects of acidity, sample flow rate and volume, elution solution, and interfering ions on recovery of the two mercury species have been systematically investigated. Under optimal conditions, the limits of detection (LODs) were 72 and 44 ng L(-1) for Hg(2+) and MeHg(+) (as Hg) with online concentration factors of 12.5 and 12.1, respectively. The relative standard deviations (RSDs) for nine replicate determinations at 5 ng mL(-1) levels of mercury species were 2.7 and 2.0% for Hg(2+) and MeHg(+), respectively. The calibration graphs were linear with a correlation coefficient of 0.9902 in the range of 0.5-100 ng mL(-1) for Hg(2+) and 0.9976 in the range of 0.1-100 ng mL(-1) for MeHg(+), respectively. The developed method was successfully applied to the direct determination of MeHg(+) and Hg(2+) in seafood samples, and the recoveries for the spiked samples were in the range of 89.9-102.4% (MeHg(+)) and 87.0-104.6% (Hg(2+)), respectively. The method was validated by analyzing a certified reference material DORM-2 (dogfish muscle), and the determined values were in good agreement with certified values.     

Study of anticancer activities of muscadine grape phenolics in vitro

Muscadine grapes have unique aroma and flavor characteristics. Although a few studies reported high polyphenols content of muscadine grapes, little research has been conducted to evaluate the phenolic compounds bioactivities in any muscadine grape cultivar. The objective of this study was to evaluate the effect of phenolic compounds in muscadine grapes on cancer cell viability and apoptosis. Four cultivars of muscadine (Carlos, Ison, Noble, and Supreme) were assessed in this study. Phenolic compounds were extracted from muscadine skins and further separated into phenolic acids, tannins, flavonols, and anthocyanins using HLB cartridge and LH20 column. Some individual phenolic acids and flavonoids were identified by HPLC. Anthocyanin fractions were more than 90% pure. The effect of different fractions on the viability and apoptosis of two colon cancer cell lines (HT-29 and Caco-2) was evaluated. A 50% inhibition of cancer cell population growth for the two cell lines was observed at concentrations of 1-7 mg/mL for crude extracts. The phenolic acid fractions showed a 50% inhibition at the level of 0.5-3 mg/mL. The greatest inhibitory activity was found in the anthocyanin fraction, with a 50% inhibition at concentrations of approximately 200 microg/mL in HT-29 and 100-300 microg/mL in Caco-2. Anthocyanin fractions also resulted in 2-4 times increase in DNA fragmentation, indicating the induction of apoptosis. These findings suggest that polyphenols from muscadine grapes may have anticancer properties.     

Liquid chromatographic determination of the estrogens daidzein, formononetin, coumestrol, and equol in bovine blood plasma and urine

A liquid chromatographic (LC) method is described for the determination of the plant estrogens diadzein, formononetin, and coumestrol and the estrogenically active metabolite equol in bovine blood plasma and urine. The blood and urine samples are incubated overnight with and without beta-glucuronidase/sulfatase for analysis of both free and conjugated forms of estrogens. Samples are applied to Extrelut columns, extracted with ethyl acetate, and evaporated to dryness. Residues from urine samples are dissolved in methanol, diluted with water, acidified with HCl, and purified by injection through a Sep-Pak C18 cartridge. This eluate is used for LC analysis. Residues from blood samples are dissolved in benzene-petroleum ether (1 + 1), extracted with ammonium hydroxide, acidified with glacial acetic acid, and extracted with ethyl acetate. The ethyl acetate extract is evaporated, dissolved in 80% methanol, injected onto a LC reverse-phase column, and separated in a linear gradient system between 40 and 80% methanol in phosphate buffer. Quantitation is performed by means of UV and fluorescence responses. The method was sensitive enough to determine 0.4 ng/mL of daidzein and formononetin and 0.1 and 13 ng/mL of coumestrol and equol, respectively, in blood, and 130, 80, and 7 ng/mL of daidzein, formononetin, and coumestrol, respectively, and 4 micrograms/mL of equol in urine. The applicability of the method was checked by the determination of total and free plant estrogens in blood samples from a dairy cow fed a normal diet.     

复混肥中缩二脲含量对作物生长的影响

采用水培和盆栽法,研究了复混肥中不同含量的缩二脲对作物生长的影响.水培施肥水平为每升水溶液中复混肥(10-8-7)含量分别为0.5 g和1.0 g,即每升水溶液中含氮量达到56.05 mg和112.1 mg;土壤施肥水平为每千克土壤中复混肥(10-8-7)含量分别为0.5 g和1.0 g,即每千克土壤中含氮量达到56.05 mg和121.1 mg.水培试验结果表明:每升水溶液中,在适宜各供试作物生长的施肥条件下,复混肥中缩二脲对冬小麦、玉米、番茄、油菜、水稻的毒害作用的临界值分别为35、40、20、15、15 mg.盆栽试验结果表明:在适宜各供试作物生长的施肥条件下,缩二脲对冬小麦、玉米、番茄、油菜、水稻的毒害作用的临界值分别为所施复混肥(10-8-7)含量的1.6%、2.0%、2.0%、2.0%、2.0%.当高于这个临界值时,缩二脲对此5种作物会造成毒害作用.考虑到复混肥的实际制造过程中大量使用含氯化肥,氯离子含量波动大,建议国家在制定复混肥产品标准时缩二脲含量应限制在1%以下为宜.     

Contribution of tomato phenolics to antioxidation and down-regulation of blood lipids

This study was performed to understand the characteristics and biological activities of phenolics in tomatoes and to examine the effect of tomato on the regulation of blood lipids. Tomatoes of both big and small sizes were used fresh, after blanching, or after blanching and heating. Moreover, a human clinical trial was conducted to examine plasma antioxidation, status of blood lipids, and phenolic responses after ingestion of fresh tomato, tomato juice, and a lycopene drink. The contents of tomato phenolics were increased by 34% for small tomato and by 23% for big tomato after treatment by blanching and heating at 100 degrees C for 30 min. Tomato phenolics showed fair antioxidant activity (57-71%) and also synergistically promoted the antioxidation (81-100%) of tomato carotenoids. In the human clinical study, total antioxidant capacity and phenolic contents in plasma were increased after administration of fresh tomato and tomato juice, but no significant difference was found for lycopene drink consumption. Triglyceride levels and low-density lipoprotein cholesterol were decreased after administration of fresh tomato and tomato juice, and high-density lipoprotein cholesterol was increased.     

Phenolic compounds from blueberries can inhibit colon cancer cell proliferation and induce apoptosis

Research has shown that diets rich in phenolic compounds may be associated with lower risks of several chronic diseases including cancer. This study systematically evaluated the bioactivities of phenolic compounds in rabbiteye blueberries and assessed their potential antiproliferation and apoptosis induction effects using two colon cancer cell lines, HT-29 and Caco-2. Polyphenols in three blueberry cultivars, Briteblue, Tifblue, and Powderblue, were extracted and freeze-dried. The extracts were further separated into phenolic acids, tannins, flavonols, and anthocyanins using an HLB cartridge and LH20 column. Some individual phenolic acids and flavonoids were identified by HPLC with >90% purity in anthocyanin fractions. The dried extracts and fractions were added to the cell culture medium to test for antiproliferation activities and induction of apoptosis. Flavonol and tannin fractions resulted in 50% inhibition of cell proliferation at concentrations of 70-100 and 50-100 microg/mL in HT-29 and Caco-2 cells, respectively. The phenolic acid fraction showed relatively lower bioactivities with 50% inhibition at approximately 1000 microg/mL. The greatest antiproliferation effect among all four fractions was from the anthocyanin fractions. Both HT-29 and Caco-2 cell growth was significantly inhibited by >50% by the anthocyanin fractions at concentrations of 15-50 microg/mL. Anthocyanin fractions also resulted in 2-7 times increases in DNA fragmentation, indicating the induction of apoptosis. The effective dosage levels are close to the reported range of anthocyanin concentrations in rat plasma. These findings suggest that blueberry intake may reduce colon cancer risk.     

Enzyme-linked immunosorbent assay for T-2 toxin metabolites in urine

A direct competitive enzyme-linked immunosorbent assay (ELISA) for determination of total T-2 toxin metabolites in urine was developed. The assay involves coating anti-3-acetyl-neosolaniol-hemisuccinate-bovine serum albumin conjugate (anti-3-Ac-NEOS-HS-BSA) antibody to the ELISA plate and using 3-Ac-NEOS-HS-peroxidase as the enzyme marker. Competitive ELISA revealed that the antibody had good cross-reactivity with acetyldiacetoxyscirpenol (Ac-DAS), T-2 tetraol tetraacetate, 3'-OH-Ac-T-2, 3-Ac-NEOS, and 3,4,15-triacetyl-12,13-epoxytrichothec-9-en-8-one (Ac-T-2-8-one), but less cross-reactivity with Ac-T-2 toxin and T-2 toxin. All metabolites of T-2 toxin in urine were converted to T-2 tetraol tetraacetate (T-2-4ol-4Ac) by acetylation of the sample extract before ELISA. To test the ELISA accuracy, a radioimmunoassay (RIA) was performed simultaneously. The linear portion of the standard curve of this direct ELISA for T-2-4ol-4Ac was 0.2-2.0 ng/mL, which was 10 times more sensitive than RIA. The minimum detection level for T-2-4ol-4Ac was 0.02 ng/mL (0.4 pg/assay) in the absence of urine sample. The overall analytical recoveries for T-2 toxin, HT-2, T-2-4ol, 3'-OH-HT-2, NEOS, and a mixture of these 5 toxins added to the urine samples in the ELISA at concentrations of 0.05 and 0.2 ng/mL were 87 and 94%, respectively.     

High performance liquid chromatographic method using fluorescence detention for quantitative analysis of zearalenone and alpha-zearalenol in blood plasma

A sensitive, high performance liquid chromatographic method is described for quantitative determination of zearalenone and alpha-zearalenol in blood plasma. Blood plasma is extracted with 2-propanol in ether, the extract is evaporated to dryness, and the residue is dissolved in 0.18N NaOH. The aqueous phase is washed with chloroform, dichloromethane, and benzene, neutralized with 0.10M H3PO4, and extracted with benzene. The extract is evaporated, dissolved in methanol, and injected onto a reverse phase column containing LiChrosorb RP-8 under the following conditions: methanol-acetonitrile-water mobile phase, fluorescence detector, excitation wavelength 236 nm, and 418 nm cut-off emission filter. The limit of detectability (twice background) is 0.5 ng standard which is equivalent to 0.6 ng standard/mL blood plasma. Linear standard curves are observed over the range of 0-35 ng of injected zearalenone and alpha-zearalenol. The recoveries from blood plasma are 76-101% in the range of 1.5-6.0 ng standard/mL blood.     

Pharmacokinetics of anthocyanins and antioxidant effects after the consumption of anthocyanin-rich acai juice and pulp (Euterpe oleracea Mart.) in human healthy volunteers

The acai berry is the fruit of the acai palm and is traditionally consumed in Brazil but has gained popularity abroad as a food and functional ingredient, yet little information exists on its health effect in humans. This study was performed as an acute four-way crossover clinical trial with acai pulp and clarified acai juice compared to applesauce and a non-antioxidant beverage as controls. Healthy volunteers (12) were dosed at 7 mL/kg of body weight after a washout phase and overnight fast, and plasma was repeatedly sampled over 12 h and urine over 24 h after consumption. Noncompartmental pharmacokinetic analysis of total anthocyanins quantified as cyanidin-3-O-glucoside showed Cmax values of 2321 and 1138 ng/L at t max times of 2.2 and 2.0 h, and AUC last values of 8568 and 3314 ng h L(-1) for pulp and juice, respectively. Nonlinear mixed effect modeling identified dose volume as a significant predictor of relative oral bioavailability in a negative nonlinear relationship for acai pulp and juice. Plasma antioxidant capacity was significantly increased by the acai pulp and applesauce. Individual increases in plasma antioxidant capacity of up to 2.3- and 3-fold for acai juice and pulp, respectively were observed. The antioxidant capacity in urine, generation of reactive oxygen species, and uric acid concentrations in plasma were not significantly altered by the treatments. Results demonstrate the absorption and antioxidant effects of anthocyanins in acai in plasma in an acute human consumption trial.     

Development of a simple method for the determination of genistein, daidzein, biochanin A, and formononetin (biochanin B) in human urine.

A simple method was developed for the determination of free and/or total isoflavones daidzein, genistein, and their respective 4'-methoxy derivatives biochanin A and formononetin (biochanin B) at low levels in human urine. A solid-phase extraction on octadecyl silica (C(18)) columns was used for the isolation of the phytoestrogens from the matrix. An extraction on a ChemElut 1010 column connected on-line to a Florisil cartridge by a Teflon stopcock was used for effective eluate purification. A mixture of dichloromethane and ethyl acetate was used for elution of the isoflavones from the columns in tandem. The isoflavones were determined as trimethylsilyl (TMS) ethers using GC/MS-SIM after separation on an HP-5MS fused silica column. TMS ethers were obtained by using BSTFA containing 1% of TMCS. For the determination of free isoflavones 6-hydroxyflavone was used as internal standard, whereas robigenin was used in the case of total isoflavone determination. Recoveries for free isoflavones under study varied from 63.5 to 89.6% at the 25 ng mL(-)(1) level and from 63.5 to 89. 2% at the 5 ng mL(-)(1) level in urine. Analytical curves were linear between 5 and 25 ng mL(-)(1). Detection limits varied from 1 ng mL(-)(1) for formononetin to 2.3 ng mL(-)(1) for daidzein. Recoveries for total isoflavone determination after enzymatic hydrolysis with glucuronidase from Helix pomatia ranged from 56.5 to 77.1% at the 25 ng mL(-1) level.     

Optimization of extraction of phenolic compounds from flax shives by pressurized low-polarity water

Pressurized low-polarity water (PLPW) extraction of phenolic compounds from flax shive was investigated using statistically based optimization and the "one-factor-at-a-time" method. Extraction variables examined using central composite design (CCD) included temperature, flow rate, and NaOH concentration of the extracting water. Extraction of phenolic compounds including p-hydroxybenzaldehyde, vanillic acid, syringic acid, vanillin, acetovanillone, and feruric acid was affected by temperature and NaOH concentration; and extraction of all phenolic compounds, except ferulic acid, increased with temperature and NaOH concentration of the extracting water. Flow rate had little effect on concentration of phenolic compounds at equilibrium, but the extraction rate at the early phase was higher for higher flow rates. The mechanism of PLPW extraction of flax shive phenolics was also investigated using a two-site kinetic model and a thermodynamic model. To determine the extraction mechanism, flow rate was varied from 0.3 to 4.0 mL/min while temperature and NaOH concentration were fixed at 180 degrees C and 0.47 M, respectively. The flow rate tests showed the extraction rates of total phenolic (TP) compounds increased with flow rate and can be described by a thermodynamic model. The results from the thermodynamic model demonstrated that a K(D) value of 30 agreed with the experimental data in the flow rate range of 0.3-4.0 mL/min. When the effect of the three independent variables was evaluated simultaneously using CCD, a maximum TP concentration of 5.8 g/kg of dry flax shive (DFS) was predicted from the combination of a high temperature (230.5 degrees C), a high initial concentration of NaOH (0.63 M), and a low flow rate (0.7 mL/min). Maximum TP concentration of 5.7 g/kg of DFS was obtained from extraction conditions of 180 degrees C, 0.3 or 0.5 mL/min, and 0.47 M NaOH at equilibrium. A second-order regression model generated by CCD predicted a maximum TP concentration of 5.8 g/kg of DFS under the same extraction conditions, which is well matched with the results from experimental data.     

Phenolic antioxidants richly contained in corn bran are slightly bioavailable in rats

Phenolic acids (PAs) have been shown to be beneficial to human health and are found most abundantly in corn bran ( approximately 4%, w/w), one of the main dietary fibers. This study therefore evaluated the bioavailabilities of phenolic antioxidants ferulic acid (FA) and p-coumaric acid (PCA) in refined corn bran (RCB) by determining their recovery in the plasma, urine, and feces of rats fed a single meal of a RCB diet containing 5% RCB or adapted to the RCB diet for 10 days. In both studies, 0.4-0.5% of ingested FA and 1.2-2.3% of ingested PCA were recovered in rat urine. By contrast, approximately 81% of FA and approximately 64% of PCA ingested with the single meal were excreted through the rat feces within 3 days after the ingestion. On the other hand, after rats were fed the RCB diet, total FA (all forms of FA) was recovered in plasma at a concentration of 35.0 +/- 2.0 microg/L, total FA and total PCA were excreted through urine at levels of 155.4 +/- 5.8 and 50.9 +/- 6.6 microg/day, respectively. These parameters showed no significant change (P = 0.93, 0.09, and 0.66, respectively) after rats were fed the RCB diet continuously for up to 10 days. These results suggest that the PAs in RCB are bioavailable in rats. Their bioavailabilities, however, are relatively low compared with their high content in RCB and not improved by the adaptation for 10 days to the enriched RCB diet. Additionally, comparison with the results of other studies revealed that high contents of FA and, especially, diferulic acids in cereal bran, which act as cross-links between bran cell wall polysaccharides, may not improve but, rather, limit the bioavailabilities of PAs in vivo.     

Chlorinated compounds and phenols in tissue, fat, and blood from rats fed industrial waste site soil extract: methods and analysis

A method was developed to analyze rat tissue, fat, and blood for some of the chlorinated compounds found in an extract of soil from an industrial waste site. Extraction with hexane and then with ethyl ether-hexane (1 + 1) was followed by concentration over steam, and gas chromatographic analysis with an electron capture detector. Volatile compounds were analyzed in a glass column coated with 6% SP-2100 plus 4% OV-11 on Chromosorb W. Semivolatile compounds, chlorinated compounds, and pesticides were analyzed in a 70 m glass capillary column coated with 5% OV-101. Phenols were analyzed in a glass column packed with 1% SP-1240 DA on Supelcoport. However, the most efficient means of separation was to use the same glass column for volatile compounds, a DB-5 fused silica capillary column for semivolatile compounds, pesticides, and phenols, and the same 1% SP-1240 DA glass column for separation of beta-BHC and pentachlorophenol. Recoveries ranged from 86.3 +/- 9.1% (mean +/- standard deviation) to 105 +/- 10.4%. Sensitivities for semivolatile chlorinated compounds, pesticides, and phenols were about 4 ng/g for fat, 1 ng/g for tissue, and 0.2 ng/mL for blood. Sensitivities for volatile compounds were about 4-fold higher (16, 4, and 0.8, respectively). Sensitivities for dichlorobenzenes and dichlorotoluenes were 8 ng/g for fat, 2 ng/g for tissue, and 0.4 ng/mL for blood.     

Evaluation of a method to characterize the phenolic profile of organic and conventional tomatoes

The present study aims to compare the phenolic profiles of organic and conventional tomatoes bought in the market. For the quantification and identification of individual polyphenols, liquid chromatography coupled to mass spectrometry in tandem mode (LC-MS/MS) was carried out. Confirmation of the compounds previously identified on the triple-quadrupole was accomplished by injection in the high-resolution system (QToF-MS). In this way, 34 compounds were identified in tomato fruits. Recoveries of targeted polyphenols exceed 78% for conventional and organic tomatoes, respectively. The method intraday precision ranged between 3 and 5%, whereas the interday one was below 12%. Comparing the analyses of tomatoes from conventional and organic production systems demonstrated statistically higher levels (P < 0.05) of phenolic compounds in organic tomatoes. This methodology allowed finding differences in the bioactive components of organic and conventional tomatoes not previously reported.