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The effect of commercial canning and freezing on the nutritional content of fresh apricots was investigated. Processed samples were analyzed post‐processing and after 3 months of storage and compared directly to fresh apricots from the same source. Vitamin C, beta‐carotene, total phenols, and antioxidants were quantified. Compared to fresh, canned apricots initially exhibited similar levels of antioxidants, a 17% increase in beta‐carotene, and a 48% increase in phenols, while vitamin C was reduced by 37%. After 3 months of storage, antioxidant levels were 47% higher than fresh. Vitamin C did not change significantly following storage and beta‐carotene decreased by 15%. The canned apricot fruit packed in light syrup did not have higher total soluble solids (TSS) levels indicating no increase in fruit sugar content. Frozen apricots exhibited large increases in antioxidants (529%), beta‐carotene (35%), vitamin C (3,370%), and phenols (406%) compared to fresh. After 3 months of storage, frozen apricots decreased in vitamin C (29%) and phenols (17%), but remained 2,375% and 318% higher than fresh, respectively. Beta‐carotene increased during storage, reaching levels 56% higher than fresh while antioxidant activity was unchanged. This study demonstrates that key nutrients in canned and frozen apricots are retained or amplified upon processing, with the exception of vitamin C in canned apricots. The routine addition of citric and ascorbic acid to fruit prior to freezing resulted in significantly higher antioxidants, vitamin C, and phenols. Consumers eating canned or frozen apricots can feel confident of similar or superior nutritional content as compared to fresh apricots. The apricot industry is limited by the short shelf life of the fruit and consumer belief that processed produce is not as nutritious as fresh. Assessing the nutritional content of canned and frozen apricots and determining that processed apricots can deliver nearly comparable nutrient levels to fresh apricots provides the evidence needed to dispel these misconceptions and potentially increase demand for processed apricots among consumers.

The effect of two passive drying methods on the degradation of ascorbic acid (vitamin C), α-tocopherol (vitamin E), and β-carotene (provitamin A) in ripe mango and tomato was determined. Samples of both commodities were dried outdoors in a solar drying cabinet or indoors in sealed plastic boxes containing ceramic zeolite drying beads in a bead-to-fruit ratio of 1:10. Nutrient content of the dried samples was compared to fresh samples (undried control) obtained from the same batch of fruit. Mango samples tended to reach an equilibrium weight of approximately 18% of their initial weight after 28 to 30 hr of drying in both drying methods, while tomatoes equilibrated at 5% to 6% of their initial weight after 8 to 10 hr of drying in the solar drying cabinet and after approximately 30 hr using drying beads. The content of all three nutrients in solar dried tomato, and of β-carotene in mango were significantly (P < 0.05) lower than the control. α-Tocopherol and β-carotene content in tomato and β-carotene content in mango were significantly lower than the control in samples dried using drying beads. Drying method had a significant (P < 0.05) effect on nutrient content; ascorbic acid and β-carotene content were lower in solar dried tomato than in tomato dried with drying beads. For mango, β-carotene content was lower after solar drying as compared to drying with drying beads. Drying fruit at lower temperatures using drying beads resulted in higher overall nutrient content of the dried fruit. PRACTICAL APPLICATION Agriculturalists in developing countries still depend largely on solar drying methods as a means of extending the shelf life and increasing the value of fruit and vegetable products. Zeolite drying beads are reusable and are capable of drying fruit relatively quickly in a controlled environment, and may reduce the degradation of certain nutrients due to heat during solar drying.

Acrylamide, formed in certain plant foods at elevated cooking temperatures, has been identified as a probable carcinogen. A wide variation in acrylamide concentration has been reported for commercial canned black ripe olives, with some levels being quite high. Standards for olive sterilization were developed before acrylamide was discovered to be present in food. The objective of this study was to determine if alternative sterilization conditions during thermal processing could substantially reduce acrylamide levels in black ripe olives, while still maintaining safety and quality. Heat penetration tests for six thermal processes were used to evaluate process F 0 – a sterilization value –which was correlated to acrylamide formation and changes in quality attributes of black ripe olives. Acrylamide concentration followed a positively correlated second order polynomial regression with process F 0 . This correlation was further demonstrated by two of the experimental processes with different initial temperatures and different processing times, but with ultimately similar F 0 , that produced very similar levels of acrylamide. The amounts of solids leaching from olives in brine increased at higher temperatures of thermal processing, whereas pH variation in olives and brine was most likely related to the differences in lye-wash cycles. Skin color was unaffected, whereas increasing (time/temperature) thermal processing reduced the firmness of whole olives. Optimization of safe thermal processing conditions while lowering process F 0 is a practical and efficient strategy to reduce acrylamide formation and improve the safety of black ripe olives.

Four vitamins were analyzed in several fruit and vegetable commodities to evaluate the differences between fresh and frozen produce. Ascorbic acid, riboflavin, α-tocopherol, and β-carotene were evaluated in corn, carrots, broccoli, spinach, peas, green beans, strawberries, and blueberries. Samples of each commodity were harvested, processed, and analyzed for nutrient content at three storage times per treatment. Ascorbic acid showed no significant difference for five of the eight commodities and was higher in frozen samples than fresh for the remaining three commodities. Apart from broccoli and peas, which were higher and lower in frozen vs fresh samples, respectively, none of the commodities showed significant differences with respect to riboflavin content. Three commodities had higher levels of α-tocopherol in the frozen samples, while the remaining commodities showed no significant difference between fresh and frozen. β-Carotene was not found in significant amounts in blueberries, strawberries, and corn. Peas, carrots, and spinach were lower in β-carotene in the frozen samples, while green beans and spinach showed no significant difference between the two storage methods. Overall, the vitamin content of the frozen commodities was comparable to and occasionally higher than that of their fresh counterparts. β-Carotene, however, was found to decrease drastically in some commodities.

Minerals, total phenolics, and fiber were analyzed in several fruit and vegetable commodities to evaluate the differences between fresh and frozen produce. Magnesium, calcium, iron, zinc, and copper were evaluated in corn, carrots, broccoli, spinach, peas, green beans, strawberries, and blueberries. Each commodity was harvested fresh and split into two batches. Half of each commodity was kept fresh, and the other half was frozen. The nutrient content was analyzed over three storage times per treatment. The retention of nutrients was highly dependent on the commodity, but the majority of the commodities showed no significant difference between fresh and frozen for all analytes (p ≤ 0.05).

This study was conducted to evaluate the effect of molinate on retinoids homeostasis in rat testis. Molinate was administrated to male Sprague-Dawley rats (200 mg kg(-1) in corn oil, ip). Retinoid measurements were made at 6, 12, 48 and 168 h time points after administration. Testis levels of retinoic acid decreased (32 %) in a statistically significant manner at the 12 and 48 h time points. However, retinol and retinaldehyde were not significantly affected by molinate. These results suggest that molinate affects retinoic acid synthesis in testis and could contribute to understanding the molecular mechanism of molinate involved testicular toxicity.

Half-lives of α-tocopherol in plasma have been reported as 2–3 d, whereas the Elgin Study required >2 y to deplete α-tocopherol, so gaps exist in our quantitative understanding of human α-tocopherol metabolism. Therefore, 6 men and 6 women aged 27 ± 6 y (mean ± SD) ingested 1.81 nmol, 3.70 kBq of [5-14CH3]-(2R, 4'R, 8'R)-α-tocopherol. The levels of 14C in blood plasma and washed RBC were monitored frequently from 0 to 460 d while the levels of 14C in urine and feces were monitored from 0 to 21 d. Total fecal elimination (fecal + metabolic fecal) was 23.24 ± 5.81% of the 14C dose, so feces over urine was the major route of elimination of the ingested [5-14CH3]-(2R, 4′R, 8′R)-α-tocopherol, consistent with prior estimates. The half-life of α-tocopherol varied in plasma and RBC according to the duration of study. The minute dose coupled with frequent monitoring over 460 d and 21 d for blood, urine, and feces ensured the [5-14CH3]-(2R, 4'R, 8'R)-α-tocopherol (the tracer) had the chance to fully mix with the endogenous [5-14CH3]-(2R, 4'R, 8'R)-α-tocopherol (the tracee). The 14C levels in neither plasma nor RBC had returned to baseline by d 460, indicating that the t1/2 of [5-CH3]-(2R, 4′R, 8′R)-α-tocopherol in human blood was longer than prior estimates.

Clomazone (trade names Cerano and Command) is a popular herbicide used on California rice fields to control aquatic weeds. Its physicochemical characteristics indicate that it will persist primarily in the water column, where microbial degradation may drive its environmental fate. The objectives were to determine microbial degradation rates and compare the metabolic products under aerobic and anaerobic conditions similar to those in California rice fields during the summer. Time-series samples were extracted and analyzed by LC/MS/MS. Metabolic profiling revealed the following clomazone-derived transitions: m/z 240 --> 125 (clomazone), m/z 242 --> 125 (ring-open clomazone), m/z 256 --> 125 (5-hydroxyclomazone), m/z 256 --> 141 (aromatic hydroxyclomazone), m/z 268 --> 125 (unknown metabolite), and m/z 272 --> 141 (4'5-dihydroxyclomazone). Results indicate an anaerobic half-life of 7.9 days, with ring-open clomazone reaching 67.4% of application at 38 days. Aerobically, clomazone degraded more slowly (t(1/2) = 47.3 days), forming mostly soil-bound residues. Thus, under summer conditions, clomazone is likely to dissipate rapidly from fields via anaerobic degradation.

Soil testing is an important component of a plant nutrient management program for farmers, home gardeners, and agricultural service personnel. Results from five commercially available colorimetric soil test kits were compared with standard laboratory analyses for pH, nitrate–nitrogen (NO3), phosphorus (P2O5), and potassium (K2O) content for Salinas clay loam soil with three cropping histories. The kits ranked in accuracy (frequency of match with analytical laboratory results) in the following sequence: La Motte Soil Test Kit, Rapitest, Quick Soiltest, Nitty-Gritty, and Soil Kit at 94%, 92%, 64%, 36%, and 33%. NO3 was most accurately determined by Rapitest and Quick Soiltest, P2O5 by Rapitest, and pH by La Motte Soil Test Kit. K2O was determined with equal accuracy by all but Soil Kit. The composition of the extractants may be an important factor affecting the accuracy of the test kit. For example, all kit extractants for K2O were composed of the same chemical and matched analytical laboratory results 82% of the time. By contrast, kits using an acid-based extractant for NO3 analysis more frequently matched the analytical laboratory results than kits using a zinc-based extractant (P ≤ 0.0001). La Motte Soil Test Kit had the largest range of pH measures, whereas Rapitest was relatively easy to use and interpret and is a practical choice for home gardeners or landscapers; both were more than 90% accurate for this soil type. Although an important limitation of commercial test kits is the approximate or categorical value of nutrient content (i.e., low, medium, high), accurate test kits can yield results quickly and economically for improved nutrient management.

An accurate method for measuring whole blood total folate using liquid chromatography with tandem mass spectrometry is described and compared to GC/MS and a chemiluminescence assay. Whole blood from normal adults (n = 15) was fortified with a [(13)C(6)]para-aminobenzoic acid (pABA) internal standard and treated with 12.1 N hydrochloric acid at 110 degrees C for 4 h to hydrolyze all folates to pABA. Contaminants in the hydrolysate were adsorbed onto a C18 SPE cartridge. The eluate containing the folate catabolite pABA was partitioned into ethyl acetate and methylesterified with trimethylsilyldiazomethane. The methyl-pABA derivatives were quantified by positive-ion atmospheric pressure chemical ionization (APCI)LC-MS/MS. An isocratic mobile phase of acetonitrile-water (70:30) (v/v) on a C18 analytical column was used with a postcolumn reagent of 0.025% formic acid. The limit of quantitation for folate was 56.6 nmol/L RBC, and the limit of detection was 22.6 nmol/L RBC. Folate levels as determined by LC-MS/MS correlated well with the chemiluminescence assay and a GC/MS method. This new LC-MS/MS method provides enhanced sample throughput (n = 36 per day) as compared to GC/MS methods. LC-MS/MS will enable accurate measurements of red blood cell (RBC) folate in nutrition surveys and clinical trials.

Proanthocyanidin (PAC) consumption has been linked to better colonic health, but PACs are poorly absorbed, making them a target for colonic metabolism. The resulting metabolites are low molecular weight and could potentially be absorbed. To understand the effects of dietary PACs it would be important to resolve the metabolic issue and link these changes to microbial population changes in a suitable model for human digestion. Here, six crossbred female pigs were fed a diet containing 1% (w/w) of MegaNatural® Gold grape seed extract (GSE) daily for 6 days. Fecal samples were analyzed by normal phase LC coupled to fluorescence detection and LC-MS/ToF. DNA was extracted from pig fecal samples and the V3/V4 region of the 16S rRNA gene was sequenced using an Illumina MiSeq. Intact parent PACs (dimer-pentamer) were observed in the feces on days 3 and 6 at similar high levels (∼400 mg kg(-1) total) during ingestion of GSE but were absent 48 h post-feeding. The major phenolic metabolites were 4-hydroxyphenylvaleric acid and 3-hydroxybenzoic acid which increased by ∼30 and 3 mg kg(-1) respectively. The GSE diet also caused an ecological shift in the microbiome, dramatically increasing Lachnospiraceae, Clostridales, Lactobacillus and Ruminococcacceae. The relationship between dietary PACs and colon health may be attributable to the altered bacterial populations or phenolic compounds in the colon.

A rapid LC-MS/MS method was developed for the quantitative determination of grayanotoxins I, II, and III in rumen contents, feces, and urine. The grayanotoxins were extracted from solid samples with methanol. The methanol extract was diluted with water and cleaned up using a reversed phase solid phase extraction column. HPLC separation was performed by reversed phase HPLC using a gradient of water and methanol containing 1% acetic acid. Determination was by positive ion electrospray ionization and ion trap tandem mass spectrometry. Grayanotoxin I quantitation was based on fragmentation of the sodium adduct ion at m/z 435 to a product ion at m/z 375. Grayanotoxins II and III were quantitated on the basis of fragmentation of the ion at m/z 335 to the product ion at m/z 299. The method detection limits were 0.2 microg/g in rumen contents and feces and 0.05 microg/g in urine. Fortifications at the detection limits and 10 times the detection limits of bovine rumen contents, caprine feces, and ovine urine were recovered in the range 80-114%. The diagnostic utility of the method was tested by analyzing samples submitted to the veterinary toxicology laboratory.

A rapid procedure was developed for the determination of nine anticoagulant rodenticides in animal serum by high-performance liquid chromatography (HPLC) with a fluorescence and photodiode array detector (DAD). The anticoagulants coumafuryl, pindone, warfarin, coumachlor, diphacinone, chlorphacinone, bromadialone, brodifacoum, and difethialone were extracted and quantitated simultaneously with this method. Anticoagulants were extracted at pH 5.5 with 5% (v/v) ethanol in ethyl acetate and 1% (w/v) trichloroacetic acid. Sample extracts were subjected to a cleanup on a Florisil solid-phase extraction (SPE) column. Separation was performed with a reversed phase amine-deactivated C18 column with a gradient of phosphate buffer, acetonitrile, and methanol. The eluent was monitored with the fluorescence detection at an excitation wavelength of 310 nm and an emission wavelength of 390 nm, and the photodiode array detection was set to 325 nm. Recoveries from spiked serum samples at ranges of 0.004−1 ppm were betwee...

Background The plant genus Fallopia is well-known in Chinese traditional medicine and includes many species that contain bioactive compounds, namely phytoestrogens. Consumption of phytoestrogens may be linked to decreased incidence of breast and prostate cancers therefore discovery of novel phytoestrogens and novel sources of phytoestrogens is of interest. Although phytoestrogen content has been analyzed in the rhizomes of various Fallopia sp., seeds of a Fallopia sp. have never been examined for phytoestrogen presence. Methods Analytical chemistry techniques were used with guidance from an in vitro estrogen receptor bioassay (a stably transfected human ovarian carcinoma cell line) to isolate and identify estrogenic components from seeds of Fallopia convolvulus. A transiently transfected human breast carcinoma cell line was used to characterize the biological activity of the isolated compounds on estrogen receptors (ER) α and β. Results Two compounds, emodin and the novel flavan-3-ol, (−)-epiafzelechin-3-O-p-coumarate (rhodoeosein), were identified to be responsible for estrogenic activity of F. convolvulus seed extract. Absolute stereochemistry of rhodoeosein was determined by 1 and 2D NMR, optical rotation and circular dichroism. Emodin was identified by HPLC/DAD, LC/MS/MS, and FT/ICR-MS. When characterizing the ER specificity in biological activity of rhodoeosein and emodin, rhodoeosein was able to exhibit a four-fold greater relative estrogenic potency (REP) in breast cells transiently-transfected with ERβ as compared to those transfected with ERα, and emodin exhibited a six-fold greater REP in ERβ-transfected breast cells. Cell type-specific differences were observed with rhodoeosein but not emodin; rhodoeosein produced superinduction of reporter gene activity in the human ovarian cell line (> 400% of maximum estradiol [E2] induction) but not in the breast cell line. Conclusion This study is the first to characterize the novel flavan-3-ol compound, rhodoeosein, and its ability to induce estrogenic activity in human cell lines. Rhodoeosein and emodin may have potential therapeutic applications as natural products activating ERβ, and further characterization of rhodoeosein is necessary to evaluate its selectivity as a cell type-specific ER agonist.

A rapid, accurate, and efficient method for determining the sterol, uvaol, and erythrodiol concentrations was developed to meet International Olive Council (IOC) certification criteria for extra virgin olive oil (EVOO). The unsaponifiable fraction of the sample (0.2 g) was separated with a diatomaceous earth column, and the sterol and triterpenic dialcohols were isolated with a novel base-activated silica solid-phase extraction (SPE) cartridge cleanup protocol. The improved method and the IOC method provided identical pass/fail results (n = 34) for each of the six sterol and erythrodiol/uvaol IOC criteria used to assess olive oil. This method was validated, and recoveries of stigmasterol (88%) and β-sitosterol (84%) were greater than previously published values obtained using the IOC method. This method requires approximately one-third the time required to complete the IOC method and has great utility for the rapid screening of EVOO to detect adulteration, false labeling, and an inferior product.

A new multiresidue analytical screen for estimating alkaloid exposure in livestock has been evaluated on biological samples from goats dosed with sublethal amounts of five plants known to cause acute poisoning in animals. The plant species selected were Conium maculatum (poison hemlock), Nicotiana glauca (tree tobacco), Delphinium barbeyi (larkspur), Datura wrightii (jimsonweed), and Taxus baccata (English yew). Animals were euthanized when toxic signs developed or when 3−7 hours had passed after the dose. The liver, kidney, rumen contents, abomasal contents, urine, and serum from each animal were examined for the presence of alkaloids. Alkaloid contents were determined by GC with nitrogen−phosphorus detection, GC/MS, and a modified commercial thin layer chromatography system. Alkaloids from the plants were detected at levels greater than 1 μg/g in samples of rumen and abomasal contents and in most urine, kidney and liver samples. No alkaloids were detected in serum samples at concentrations greater than ...

A high-performance liquid chromatography (HPLC)-fluorescence method was developed for quantitative determination of oleandrin in gastrointestinal contents (stomach, rumen, colon, and cecum contents). Oleandrin was extracted with methylene chloride. The extract was cleaned up on a charcoal solid phase extraction (SPE) column, derivatized with 1-naphthoyl chloride, and quantified by HPLC with a fluorescence detector. The limit of detection was 0.05 ppm. Six replicate fortifications of stomach contents matrix at 0.10 ppm oleandrin gave an average recovery of 85%, with 4.6% CV (relative standard deviation). The diagnostic utility of the method was tested by analyzing samples submitted to the veterinary toxicology laboratory.

A selective and sensitive method was developed for the quantitative determination of pyrogallol and detection of gallic acid in biological samples. Pyrogallol was measured as a metabolite of gallotannin in bovine urine and serum samples. Gallic acid was formed by hydrolysis of the gallotannins in blue oak (Quercus douglasii) leaves, tannic acid standards, and rumen contents. Acid hydrolyzed or acidified samples were extracted with 5% ethanol in ethyl acetate (v/v), followed by evaporation and derivatization with Deriva-sil. The trimethylsilyl derivatives were analyzed by gas chromatography/mass spectrometry using selected ion monitoring. Method detection limits for gallic acid and pyrogallol were 0.5 ppm in urine and serum samples and 5 ppm in rumen contents. The diagnostic utility of the method was tested by analyzing samples from heifers dosed with blue oak leaves and commercial tannic acid.

Kinetic models enable nutrient needs and kinetic behaviors to be quantified and provide mechanistic insights into metabolism. Therefore, we modeled and quantified the kinetics, bioavailability, and metabolism of RRR-α-tocopherol in 12 healthy adults. Six men and 6 women, aged 27 ± 6 y, each ingested 1.81 nmol of [5(-14)CH(3)]-(2R, 4'R, 8'R)-α-tocopherol; each dose had 3.70 kBq of (14)C. Complete collections of urine and feces were made over the first 21 d from dosing. Serial blood samples were drawn over the first 70 d from dosing. All specimens were analyzed for RRR-α-tocopherol. Specimens were also analyzed for (14)C using accelerator MS. From these data, we modeled and quantified the kinetics of RRR-α-tocopherol in vivo in humans. The model had 11 compartments, 3 delay compartments, and reservoirs for urine and feces. Bioavailability of RRR-α-tocopherol was 81 ± 1%. The model estimated residence time and half-life of the slowest turning-over compartment of α-tocopherol (adipose tissue) at 499 ± 702 d and 184 ± 48 d, respectively. The total body store of RRR-α-tocopherol was 25,900 ± 6=220 μmol (11 ± 3 g) and we calculated the adipose tissue level to be 1.53 μmol/g (657 μg/g). We found that a daily intake of 9.2 μmol (4 mg) of RRR-α-tocopherol maintained plasma RRR-α-tocopherol concentrations at 23 μmol/L. These findings suggest that the dietary requirement for vitamin E may be less than that currently recommended and these results will be important for future updates of intake recommendations.

Using linear regression models, we studied the main and 2-way interaction effects of the predictor variables gender, age, BMI, and 64 folate/vitamin B-12/homocysteine (Hcy)/lipid/cholesterol-related single nucleotide polymorphisms (SNP) on log-transformed plasma Hcy normalized by RBC folate measurements (nHcy) in 373 healthy Caucasian adults (50% women). Variable selection was conducted by stepwise Akaike information criterion or least angle regression and both methods led to the same final model. Significant predictors (where P values were adjusted for false discovery rate) included type of blood sample [whole blood (WB) vs. plasma-depleted WB; P < 0.001] used for folate analysis, gender (P < 0.001), and SNP in genes SPTLC1 (rs11790991; P = 0.040), CRBP2 (rs2118981; P < 0.001), BHMT (rs3733890; P = 0.019), and CETP (rs5882; P = 0.017). Significant 2-way interaction effects included gender × MTHFR (rs1801131; P = 0.012), gender × CRBP2 (rs2118981; P = 0.011), and gender × SCARB1 (rs83882; P = 0.003). The relation of nHcy concentrations with the significant SNP (SPTLC1, BHMT, CETP, CRBP2, MTHFR, and SCARB1) is of interest, especially because we surveyed the main and interaction effects in healthy adults, but it is an important area for future study. As discussed, understanding Hcy and genetic regulation is important, because Hcy may be related to inflammation, obesity, cardiovascular disease, and diabetes mellitus. We conclude that gender and SNP significantly affect nHcy.

A multiresidue screen for the quantitative determination of 43 organ op hosphorus, 17 orga no-chlorine, and 11 N-methyl carbamate insecticides in 10 g of plant or animal tissues is described. The insecticides are extracted with 5% ethanol in ethyl acetate (v/v). Samples with high lipid content are cleaned up by automated gel permeation chromatography with a 30% ethyl acetate in hexane (v/v) eluant and in-line silica gel minicolumns. Highly pigmented samples are cleaned up with class-specific solid-phase extraction columns. The concentrated extracts are analyzed by selective detection with gas chromatography or liquid chromatography. Recovery of 71 insecticides ranged from 77 to 113%. Analysis of fortified bovine liver (n = 5) resulted in an average recovery of 96 ± 4% at the 0.5 to 0.05 μg/g level. Analysis of fortified alfalfa hay (n = 5) resulted in a mean recovery of 94 ± 4% at the 0.06 to 0.5 μg/g level, and analysis of fortified fresh tomatoes (n = 5) resulted in an average recovery of 97 ± 3% at the 0.06 to 0.5 μg/g level. Method detection limits ranged from 0.02 to 0.5 μg/g for the compounds studied with a nominal 10 g sample.

A rapid method is described for the quantitative determination of cholinesterase activity in large batches of blood and brain samples. The technique is an adaptation of the Ellman procedure for a 96-well microtiter plate reader. Ten samples can be analyzed simultaneously in 5 min, with all calculations, including statistical analysis, done automat- ically. The method detection limit is 0.1 μM/mL/min for blood and 0.1 μM/g/min for brain samples. The procedure has been applied to the routine analyses of samples presented to the veterinary diagnostic laboratory. Method performance, quality control, and normal ranges of cholinesterase activity in live- stock and other animals are described.

Half-lives of α-tocopherol in plasma have been reported as 2-3 d, whereas the Elgin Study required2 y to deplete α-tocopherol, so gaps exist in our quantitative understanding of human α-tocopherol metabolism. Therefore, 6 men and 6 women aged 27 ± 6 y (mean ± SD) ingested 1.81 nmol, 3.70 kBq of [5-(14)CH(3)]-(2R, 4'R, 8'R)-α-tocopherol. The levels of (14)C in blood plasma and washed RBC were monitored frequently from 0 to 460 d while the levels of (14)C in urine and feces were monitored from 0 to 21 d. Total fecal elimination (fecal + metabolic fecal) was 23.24 ± 5.81% of the (14)C dose, so feces over urine was the major route of elimination of the ingested [5-(14)CH(3)]-(2R, 4'R, 8'R)-α-tocopherol, consistent with prior estimates. The half-life of α-tocopherol varied in plasma and RBC according to the duration of study. The minute dose coupled with frequent monitoring over 460 d and 21 d for blood, urine, and feces ensured the [5-(14)CH(3)]-(2R, 4'R, 8'R)-α-tocopherol (the tracer) had the chance to fully mix with the endogenous [5-(14)CH(3)]-(2R, 4'R, 8'R)-α-tocopherol (the tracee). The (14)C levels in neither plasma nor RBC had returned to baseline by d 460, indicating that the t(1/2) of [5-CH(3)]-(2R, 4'R, 8'R)-α-tocopherol in human blood was longer than prior estimates.

The microbial degradation of etofenprox, an ether pyrethroid, was characterized under anaerobic (flooded) and aerobic (nonflooded) California rice field soil conditions by determination of its half-life (t1/2) and dissipation rate constant (k) and identification and quantification of degradation products at both 22 and 40 °C using LC-MS/MS. The overall anaerobic t1/2 at 22 °C ranged from 49.1 to 100 days (k=-0.0141 to -0.0069 days(-1)) compared to 27.0 days (k=-0.0257 days(-1)) at 40 °C, whereas under aerobic conditions the overall t1/2 was 27.5 days (k=-0.0252 days(-1)) at 22 °C compared to 10.1-26.5 days (k=-0.0686 to -0.0262 days(-1)) at 40 °C. The biphasic dissipation profiles were also fit to a first-order model to determine the t1/2 and k for both the fast and slow kinetic regions of the dissipation curves. Hydroxylation at the 4'-position of the phenoxy phenyl ring was the major metabolic process under anaerobic conditions for both 22 °C (maximum% yield of applied etofenprox mass=1.3±0.7%) and 40 °C (max % yield=1.2±0.8%). Oxidation of the ether moiety to the ester was the major metabolite under aerobic conditions at 22 °C (max% yield=0.5±0.1%), but at 40 °C increased amounts of the hydroxylated form were produced (max% yield=0.7±0.2%, compared to 0.3±0.1% for the ester). The hydrolytic product of the ester, 3-phenoxybenzoic acid (3-PBA), was not detected in any samples. Sterilized control soils showed little etofenprox degradation over the 56-day incubation period. Thus, the microbial population in a flooded soil was able to transform and contribute to the overall dissipation of etofenprox. The simulated summer temperature extreme (40 °C) increased the overall degradation.

It is well understood that water hemlock tubers are highly toxic to animals and to humans. However, this is the first time that immature seed from (Cicuta maculata) has been implicated in livestock poisoning. Nine mature Hereford cows from a herd of 81 died in northwestern Utah after ingesting immature seed heads of water hemlock (Cicuta maculata) in late summer. No obvious signs of poisoning were reported as all nine were found dead near the banks of the stream where water hemlock was found. Upon discovery of the dead cows, the remaining 72 cows were immediately removed from the pasture and no further losses occurred. Field necropsy of 3 of the dead cows and follow-up serology and histopathological examination of selected tissues did not identify any bacterial or viral causes. History of ingestion of large quantities of water hemlock seed, the acute nature of the deaths, chemical comparison of seed with toxic tubers and follow-up mouse bioassay testing supported the diagnosis of water hemlock poisoning. Seed heads collected from the neighboring pasture upstream and across the fence from the poisoned cattle and tubers collected from grazed plants were chemically analyzed and found to contain cicutoxin, and high levels of two cicutol-like derivatives (cicutol-#1 and #2) as well as other unidentified polyacetylene compounds. Seeds and tubers from suspected plants were semi-quantified and compared to archive samples of highly toxic tubers used in previous experiments. The immature hemlock seed contained less cicutoxin (0.01 times), but 9.5 and 22.5 times more cicutol-#1 and cicutol-#2 respectively, compared to the archive sample. Tubers from the grazed plants contained 4.6 times more cicutoxin and 9.8 and 18.8 times more cicutol-#1 and cicutol-#2 respectively, compared to the archive sample. Mouse bioassays with water extracts of immature seed and tubers from grazed plants demonstrated both were highly toxic and of greater toxicity when compared to archived sample.

A multiresidue screen for quantitative determination of 43 organophosphorus insecticides In 5 g of plant and animal tissues Is described. The organophosphorus insecticides are extracted with methanol-dichloromethane (10 + 90, v/v) and cleaned up using automated gel permeation chromatography with hexane-ethyl acetate (60 + 40) eluant and in-line silica gel minicolumns. Concentrated extracts are analyzed by gas chromatography with flame photometric detection. The method recovers 43 organophosphorus insecticides in the range of 72 to 115%. Analysis of fortified bovine liver (n = 5) shows an average 95.9 ± 4.8% recovery at the 0.05 μg/g level and 93 ± 3.8% at the 0.5 μg/g level. Analysis of fortified bovine rumen content (n = 5) shows an average 98 ± 4.2% recovery at the 0.1 μg/g level and 98.7 ± 2.8% at the 1 μg/g level. Method detection limits ranged from 0.01 to 0.05 μg/g for the compounds studied using a nominal 5 gram sample.

Previous lab studies implicated the sulfoxidation pathway of molinate metabolism to induce testicular toxicity. Once molinate is metabolized to molinate sulfoxide, it undergoes further phase II metabolism either spontaneously, enzyme catalyzed, or both to form glutathione-conjugated molinate. This study compared the metabolic capability of rat and human liver cytosol to form a glutathione (GSH)-conjugated metabolite of molinate. The GSH conjugation of molinate sulfoxide in rat cytosol was described by the constants Km of 305 microM and Vmax of 4.21 nmol/min/mg cytosol whereas the human values were 91 microM and 0.32 nmol/min/mg protein for Km and Vmax, respectively. At the same 1 mM GSH concentration, the in vitro bimolecular nonenzymatic rate constant of 3.02 x 10(-6) microM(-1) min(-1) was calculated for GSH conjugation of molinate sulfoxide. Specific activity for rat and human glutathione transferase was calculated to equal 1.202 +/- 0.25 and 0.809 +/- 0.45 micromol/min/mg protein, respectively by 1-chloro-2,4-dinitrobenzene (CDNB) assay. Compared to a conventional GSH depletion model (BSO + DEM combination), molinate alone was nearly as effective in reducing GSH levels by approximately 90 and 25% in liver and testes, respectively. The impact of molinate sulfoxide's ability to adduct glutathione transferase and inhibit the production of the glutathione conjugated metabolite was examined and found to be negligible.

An optimal folate nutritional status is important in minimizing developmental and degenerative disease. Therefore, constant monitoring of folate intake and of biomarkers of folate nutritional status is essential. The objective of this research was to compare two folate intake instruments and validate each one against RBC folate measured by a high-throughput liquid chromatography tandem mass spectrometry (HT LC-MS/MS) method described in the companion paper (Owens, J. E.; Holstege, D. M.; Clifford, A. J. J. Agric. Food Chem. 2007, 55, 3292-3297). A food frequency questionnaire (FFQ) and a folate-targeted semiquantitative Block dietary folate equivalents (DFE) screener were compared and individually validated against an HT LC-MS/MS method. RBC folate was 1178 +/- 259 nmol/L (mean +/- SD) in a population of 337 normal adult subjects. Folate intakes were 556 +/- 265 microg/day by the FFQ and 524 +/- 276 microg/day by the DFE screener. Folate intakes by the DFE screener were approximately 34 microg less than by the FFQ (paired t test, p0.01), but the intake instruments were highly correlated for total folate intake (r=0.608, p0.01). Correlations between instruments and RBC folate were low (r0.35) but strong (p0.01). ROC curve analysis indicates that the measurement of RBC folate by the HT LC-MS/MS method is a better predictive tool than are intake instruments for the evaluation of marginal folate status.

A high-throughput liquid chromatography tandem mass spectrometry (HT LC-MS/MS) method for red blood cell (RBC) folate analysis was developed from a previously described manual (M) LC-MS/MS method. The HT LC-MS/MS method used 96-well plates in which RBC folates were hydrolyzed with concentrated HCl in the presence of the [13C6]pABA internal standard (IS). The pH of the hydrolysate was adjusted to 5.0 before cleanup using 96-well plate OASIS HLB SPE cartridges. The analyte and IS were eluted with ethyl acetate/hexane (95:5, v/v) and methylated with methanol and trimethylsilyldiazomethane. The methylated analyte and IS were quantified with LC-MS/MS as previously described. The HT LC-MS/MS method was validated by determining the recovery of six different folate vitamers, which were quantitatively recovered (84-105% with CV

Milk is typically screened for beta-lactam antibiotics by nonspecific methods. Although these methods are rapid and sensitive, they are not quantitative and can yield false positive findings. A sensitive and specific method for the quantitation and mass spectral confirmation of five beta-lactam and two cephalosporin antibiotics commonly or potentially used in the dairy industry is described using high-performance liquid chromatography with tandem mass spectrometry. The antibiotics studied were ampicillin, amoxicillin, penicillin G, penicillin V, cloxacillin, cephapirin, and ceftiofur. The antibiotics were extracted from milk with acetonitrile, followed by reversed-phase column cleanup. The extract was analyzed by liquid chromatography coupled with a mass spectrometer, using a water/methanol gradient containing 1% acetic acid on a C-18 reversed-phase column. Determination was by positive ion electrospray ionization and ion trap tandem mass spectrometry. Quantitation was based on the most abundant product ions from fragmentation of the protonated ion for amoxicillin, cephapirin, ampicillin, and ceftiofur and on the fragmentation of the sodium adduct for penicillin G, penicillin V, and cloxacillin. The method was validated at the U.S. FDA tolerance or safe level and at 5 or 2.5 ng/mL for these compounds in bovine milk. Theoretical method detection limits in milk based on a 10:1 signal to noise ratio were 0.2 ng/mL (ampicillin), 0.4 ng/mL (ceftiofur), 0.8 ng/mL (cephapirin), 1 ng/mL (amoxicillin and penicillin G), and 2 ng/mL (cloxacillin and penicillin V) using a nominal sample size of 5 mL.

Six Nubian goats were exposed to an azalea branch (Rhododendron indica) at the Riverbanks Zoological Park in Columbia, South Carolina. The following day, 3 of the 6 goats were referred to the veterinary hospital of the Riverbanks Zoological Park with bloat, profuseregurgitation, and signs of depression, intermittent head pressing, and fine muscle tremors in the hind limbs. The goats were treated with magnesium hydroxide, activated charcoal, and lactated Ringer's solution and recovered within 24 hours. Definitive diagnosis of grayanotoxin exposure was accomplished by use of liquid chromatography-mass spectrometry analysis of urine and fecal samples. Rhododendron spp are members of the Ericaceae (Heath) family and may contain grayanotoxins, which exert toxic effects by binding to sodium channels in cell membranes and increasing the permeability of sodium ions in excitable membranes. Rhododendron poisoning should be considered in animals with clinical evidence of gastrointestinal tract irritation, cardiac arrhythmias, and neurologic signs.

A two-dimensional thin-layer chromatographic method was developed for the qualitative determination of the cardiotoxins oleandrin, gitoxin, digitoxin, gitoxigenin, and grayanotoxins I, II, and III in gastrointestinal contents (stomach, rumen, colon, and cecum contents), feces, and plant material. The cardiotoxins were extracted with dichloromethane. The extract was cleaned up by charcoal and reverse phase solid-phase extraction columns. Analysis was performed by two-dimensional thin-layer chromatography on silica gel plates and visualized by aluminum chloride followed by chloramine T spray. The method detection limits were 0.05 microg/g for oleandrin, 0.1 microg/g for gitoxin, and 0.2 microg/g for the other toxicants in gastrointestinal contents and feces and were 5 times higher in plant material. Four replicate fortifications of bovine rumen contents, bovine feces, and alfalfa at these levels were all well recovered. The diagnostic utility of the method was tested by analyzing samples submitted to the veterinary toxicology laboratory.

Twenty-four of 175 heifers died after ingesting water from a stock pond containing blue-green algae (genus Microcystis) in southern Colorado. Affected cattle were found dead or had signs of nervousness, and were recumbent, weak, anorectic, and hypersensitive to noise when first examined. All cattle died within 3 days after the onset of signs. At necropsy, the rumen contained blue-green algae, and the liver was larger than normal, friable, and dark red. The most important histologic lesion was hepatocyte degeneration and necrosis. Intraperitoneal administration of lyophilized cell material from the bloom caused hepatic necrosis and death in mice, and water from the pond had clumps of cells surrounded by a clear calyx, consistent with the appearance of organisms of the genus Microcystis. Samples of pond water were examined by means of high-pressure liquid chromatography; microcystin-LR, one of the hepatotoxins produced by Microcystis spp, was found. Chromatography may be useful in the diagnosis of blue-green algae toxicosis.

Eight of 600 Holstein heifers and cows died after ingestion of sweet clover silage (Melilotus sp) that contained excessive concentrations of dicumarol caused by mold infestation. The cattle developed subcutaneous hemorrhages and bled from the vagina, became weak, were unable to move, and died. To the best of our knowledge, this is the first report of sweet clover poisoning in cattle from California and is discussed in light of previous findings in the Midwest and Canada. Sweet clover poisoning is caused by dicumarol, a fungal metabolite produced from substrates in sweet clover, and is a common livestock problem in the Northern Plains and Canada. Sweet clover poisoning should be considered in livestock animals with clinical evidence of hemostatic dysfunction, prolonged coagulation times, subcutaneous hemorrhages, and hemorrhagic abortions. Definite diagnosis of moldy sweet clover poisoning can be accomplished by analysis of serum and feed samples for dicumarol concentrations.

Since mid- 1989, 37 cases of oleander poisoning in livestock have been diagnosed at the California Veterinary Diagnostic Laboratory System. The most frequent source for oleander exposure was plant clippings. Sudden death was the most common presenting complaint. Other signs reported included diarrhea, pulmonary edema, tachycardia, cardiac arrhythmias, colic, and lethargy. In the past, a presumptive diagnosis of oleander poisoning could be based only on matching clinical signs with evidence of consumption of oleander. A new 2 dimensional Thin-layer chromatography analysis of ingesta for oleandrin and an awareness of lesions in heart muscle have greatly improved the ability to diagnose oleander toxicosis.

A multiresidue screen for the quantitative determination of 43 organophosphorus, 17 organochlorine, and 11 N-methyl carbamate insecticides in 10 g of plant or animal tissues is described. The insecticides are extracted with 5% ethanol in ethyl acetate (v/v). Samples with high lipid content are cleaned up by automated gel permeation chromatography with a 30% ethyl acetate in hexane (v/v) eluant and in-line silica gel minicolumns. Highly pigmented samples are cleaned up with class-specific solid-phase extraction columns. The concentrated extracts are analyzed by selective detection with gas chromatography or liquid chromatography. Recovery of 71 insecticides ranged from 77 to 113%. Analysis of fortified bovine liver (n = 5) resulted in an average recovery of 96 +/- 4% at the 0.5 to 0.05 micrograms/g level. Analysis of fortified alfalfa hay (n = 5) resulted in a mean recovery of 94 +/- 4% at the 0.06 to 0.5 micrograms/g level, and analysis of fortified fresh tomatoes (n = 5) resulted in an average recovery of 97 +/- 3% at the 0.06 to 0.5 micrograms/g level. Method detection limits ranged from 0.02 to 0.5 micrograms/g for the compounds studied with a nominal 10 g sample.

A rapid method is described for the quantitative determination of cholinesterase activity in large batches of blood and brain samples. The technique is an adaptation of the Ellman procedure for a 96-well microtiter plate reader. Ten samples can be analyzed simultaneously in 5 min, with all calculations, including statistical analysis, done automatically. The method detection limit is 0.1 microM/mL/min for blood and 0.1 microM/g/min for brain samples. The procedure has been applied to the routine analyses of samples presented to the veterinary diagnostic laboratory. Method performance, quality control, and normal ranges of cholinesterase activity in livestock and other animals are described.

The classical mouse uterine bioassay was evaluated and adapted for routine diagnostic use in response to requests for evaluation of forages suspected of being estrogenic. Forages were extracted in acetone or 10% ethanol in acetone (v/v). Extracts were mixed with ground corn-based mouse feed. Immature female mice (n = 3/group) were fed a total of 100 g of the ground feed for 5 days. Body weights were monitored before and after the trial. After 6 days, the mice were euthanized and uterine weights were determined. Mean uterine weights were compared using 1 -way analysis of variance with preselected contrasts for individual means. Selected uteruses were fixed in 10% neutral buffered formalin for histologic examination. Control feeds, diethylstilbestrol (DES), estradiol, coumestrol, feeds with no reported estrogenic properties, and a feed that caused hyperestrogenism in cattle were tested. Moderate levels of estrogenic compounds resulted in dose-responsive uterine enlargements (10-270 ppm coumestrol over 5 days). Extremely high levels of estrogen frequently resulted in feed refusal and lack of uterine enlargement (10 ppm DES, 100 ppm estradiol). Diagnostically significant estrogenic activity was recovered from the feed known to have been estrogenic in cattle. The classical mouse uterine bioassay was relatively inexpensive, quick, repeatable, and capable of detecting clinically relevant coumestrol levels in hay.

rent veterinary therapy 7: small animal pratice, ed. Kirk RW, pp. 141-144. WB Saunders Co., Philadelphia, PA. 8. Miller S, Bauk TJ: 1992, Lead toxicosis in a group of cats. J Vet Diagn Invest 4:362-363. 9. Morgan RV, Moore FM, Pearce LK, Rossi T: 1991, Clinical and laboratory findings in small companion animals with lead poisoning: 347 cases (1977-1986). J Am Vet Med Assoc 199: 93-97. 10. Mount ME: 1989, Toxicology. In: Textbook of veterinary internal medicine, ed. Ettinger SJ, pp. 469-470. WB Saunders Co., Philadelphia, PA. 11. Prasse KW, Mahaffey EA: 1987, The hematopoietic system. In: Diseases of the cat: medicine and surgery, ed. Holzworth J, p. 764. WB Saunders Co., Philadelphia, PA. 12. Prescott CW: 1993, Clinical findings in dogs and cats with lead poisoning. Aust Vet J 60:270-271. 13. Puls R: 1988, Lead-cat tissue levels. In: Mineral levels in animal health: diagnostic data, ed. Puls R, p. 117. Sherpa International, Clearbrook, BC, Canada. 14. Reid FM, Oehme GW: 1989, Toxicoses. In: The cat-diseases and clinical management, ed. Sherding RG, p. 206. Churchill Livingstone, New York, NY. 15. Turner AJ, Fairbum AJ: 1979, Lead poisoning in the cat. Aust Vet Pract 9:205-207. 16. Watson ADJ: 1981, Lead poisoning in a cat. J Small Anim Pract 22:85-89. 17. Zook BC, Carpenter JL, Leeds EB: 1969, Lead poisoning in dogs. J Am Vet Med Assoc 155:1329-1342.

A multiresidue screen for quantitative determination of 43 organophosphorus insecticides in 5 g of plant and animal tissues is described. The organophosphorus insecticides are extracted with methanol-dichloromethane (10 + 90, v/v) and cleaned up using automated gel permeation chromatography with hexane-ethyl acetate (60 + 40) eluant and in-line silica gel minicolumns. Concentrated extracts are analyzed by gas chromatography with flame photometric detection. The method recovers 43 organophosphorus insecticides in the range of 72 to 115%. Analysis of fortified bovine liver (n = 5) shows an average 95.9 +/- 4.8% recovery at the 0.05 micrograms/g level and 93 +/- 3.8% at the 0.5 micrograms/g level. Analysis of fortified bovine rumen content (n = 5) shows an average 98 +/- 4.2% recovery at the 0.1 micrograms/g level and 98.7 +/- 2.8% at the 1 micrograms/g level. Method detection limits ranged from 0.01 to 0.05 micrograms/g for the compounds studied using a nominal 5 gram sample.

A commercial flea and tick product containing 9.0% fenvalerate for use in dogs and cats was suspected of causing illness. An acute toxicity study was performed in 10 dogs and 10 cats exposed to the product orally (po) and dermally at differing doses. Samples were obtained for DEET and fenvalerate analysis. Oral dosing of dogs and cats produced severe clinical illness at doses as low as 0.66% of a can (7 ounce spray can)/kg body weight. Dermal application of the product resulted in minor clinical abnormalities in dogs. Oral exposure at 0.5% can/kg body weight resulted in severe illness, and dermal application caused severe illness or death in cats at 20% and 40% of a can/kg body weight. The cats receiving 10% of a can/kg body weight dermally became depressed for several hours but recovered uneventfully. Serum DEET concentrations closely paralleled the clinical signs observed in the animals. Serum concentrations of DEET above 20 ppm were considered diagnostic for intoxication. Urine concentrations of DEET above 1 ppm and tissue (liver, bile, and kidney) concentrations of DEET above 10 ppm were supportive of poisoning; values near 100 ppm were diagnostic for fatal poisoning.