Gas-liquid chromatographic-mass spectrometric determination of alpha- and beta-naphthylamines in FD&C Red No. 2 (amaranth)

A method is described for the simultaneous quantitation of trace amounts of alpha- (alpha-NA) and beta-naphthylamines (beta-NA) with detectability in the 0.1 ppb range and sensitivity of 50 picomoles in certified food grade amaranth (FD&C Red No. 2; C.I. Food Red 9; CI 16185). The amaranth sample is extracted with benzene, and the evoporated residue is derivatized with perfluorooctanoic anhydride. The resulting derivatives are separated by gas-liquid chromatography and identified and quantitated by mass spectrometric monitoring of the m/e at 539.04. The method was used for quantitation of alpha-NA and beta-NA in randomly chosen samples of amaranth. Of 11 samples from different manufacturers, 5 were free of the beta-isomer; the remaining samples contained up to 1.2 ppb beta-NA. The concentration of alpha-NA ranged from no detectable amount to 970 ppb; the majority of the samples contained less than 7 ppb.     

Gas chromatographic method for ethylene dibromide in grains and grain-based products: collaborative study

Nine laboratories analyzed samples of whole grain, intermediate, and ready-to-eat products for ethylene dibromide (EDB) residues. Supplied samples of wheat, rice, and flour contained both fortified and incurred EDB; corn bread mix, baby cereal, and bread contained only fortified EDB. The whole grains and intermediates were analyzed by the same basic procedural steps as in the official method for multifumigants: They were extracted by soaking in acetone-water (5 + 1). The baby cereal and bread were analyzed by a modification of the Rains and Holder hexane co-distillation procedure. EDB was determined by electron capture gas chromatography operated with an SP-1000 column. All products contained 3 different levels of EDB and were analyzed as blind duplicates. Overall mean recoveries ranged from 85.2% for 69.6 ppb to 105.0% for 4.35 ppb, both in baby cereal. Interlaboratory relative standard deviations ranged from 5.7% for 869 ppb in wheat to 20.2% for 69.6 ppb in baby cereal, both fortified. Mean levels of incurred EDB in wheat, rice, and flour were 926.7, 982.0, and 49.9 ppb, respectively; corresponding relative standard deviations were 9.9, 7.7, and 13.1%. The method was adopted official first action.     

Fluoroacetate residues in ground squirrel and coyote tissues due to primary or secondary 1080 poisoning

Fluoroacetate residues in various tissues of 1080-poisoned ground squirrels and coyotes are listed. The tissues (excluding the stomach) of squirrels poisoned with an average of 0.8 mg 1080/kg (low dose) contained from 182 to 1309 ppb fluoroacetate. In squirrels poisoned with an average of 4.8 mg 1080/kg (high dose), the tissue residues ranged from 535 to 9754 ppb fluoroacetate. Tissues from coyotes which died after consuming 1080-poisoned ground squirrels were also analyzed for fluoroacetate residues. Residues in these coyote kidneys and livers ranged from less than 10 ppb to 95 ppb fluoroacetate. The residue findings in this research indicate that a diagnostic assay for 1080 in tissues must be reliable at 10 ppb (or less) fluoroacetate.     

Gas chromatographic determination of incurred dimetridazole residues in swine tissues

A gas chromatographic method for determination of 2-hydroxymethyl-1-methyl-5-nitroimidazole (DMZOH), the hydroxy metabolite of dimetridazole, in swime muscle has been developed. The method uses cleanup steps similar to those of an earlier polarographic method. The present method is capable of quantitating levels as low as 2 ppb and detecting less than 1 ppb. Recoveries from 30 control tissues spiked at 1, 2, or 4 ppb averaged 80.4%. Performance of the method in incurred tissue was documented and limited data on the depletion of the metabolite in muscle were generated. The muscle of swine given 150 ppm dimetridazole in feed for 14 days contained less than 1 ppb DMZOH at 12 h withdrawal time.     

浣熊黄曲霉毒素中毒病例报告

1989年10月,广西某动物园饲养的浣熊突然发病,经临床检查,出现中毒性症状:死亡后剖检和组织学检查,所见病理变化为中毒性病变;细菌学检查未见病原性细菌;已停用的饲料玉米粒含 AFT B 640 ppb;正在用的饲料玉米粉含 AFT B 235 ppb;用饲料加工的饲用混合饭团含 AFT B 184 ppb;死亡浣熊肝脏、肾脏残留的 AFT B_1分别为10 ppb 和2 ppb.诊断:浣熊是由于黄曲霉毒素亚急性中毒引起发病死亡.     

Purge and trap method for determination of ethylene dibromide in whole grains, milled grain products, intermediate grain-based foods, and animal feeds

An improved method has been developed for the determination of ethylene dibromide (EDB; 1,2-dibromoethane) in whole grains, milled grain products, intermediate grain-based foods, and animal feeds. Samples are mixed with water and sparged with nitrogen for 1 h with stirring in a water bath at 100 degrees C. The EDB collected on the adsorbent Tenax TA is eluted with hexane and determined by gas chromatography (GC) with electron capture detection (ECD) and confirmed with Hall electrolytic conductivity detection (HECD) using a second GC column. The highest levels of EDB were also confirmed by full scan GC/mass spectrometry (GC/MS). A total of 24 whole grains, milled grain products, intermediate grain-based foods, and animal feeds analyzed by using this method contained EDB levels up to 840 ppb (wheat). Recoveries from fortified samples ranged from 90 to 105%. Values from this method were compared with those obtained from the acetone soak method; for all 24 samples, this purge and trap method gave equivalent or superior recoveries and detected levels of EDB. Chromatograms for this purge and trap method were clean, enabling a quantitation level of 0.5 ppb to be achieved.     

Survey of U.S. wheat for ochratoxin and aflatoxin.

A total of 291 hard red winter wheat samples, 286 hard red spring wheat samples, and 271 soft red winter wheat samples were analyzed for the presecne of ochratoxin and aflatoxin. Samples in all grades came from those collected during crop years 1970-1973 for grade determinations by the Agricultural Marketing Service, U.S. Department of Agriculture. Sensitivity limits of the analytical method as carried out were 1-3 ppb aflatoxin B1 and 15-30 ppb ochratoxin A. No aflatoxin was detected in any sample. Three samples of hard red winter wheat (Grades U.S. No. 4 and 5 and Sample Grade) contained ochratoxin A (trace, 35, and 25 ppb, respectively). Eight of the hard red spring wheats contained ochratoxin A (15-115 PPB); these were in Grades U.S. No. 4 and 5 and Sample Grade.     

Antimutagenic effect of fruit and vegetable ethanolic extracts against N-nitrosamines evaluated by the Ames test.

The inhibitory effect of nine fruit and vegetable ethanolic extracts against the mutagenicity of N-nitrosodimethylamine (NDMA), N-nitrosopyrrolidine (NPYR), N-nitrosodibutylamine (NDBA), and N-nitrosopiperidine (NPIP) was evaluated by means of the Ames test. Licorice ethanolic extract was the only one that showed an inhibitory effect (ranging from moderate to strong) against mutagenicity of all N-nitrosamines tested. This ethanolic extract showed the greatest inhibition effect against NPIP (72%), NDMA (45%), and NPYR (39%). The greatest inhibition effect (51%) of the mutagenicity of NDBA was shown by kiwi ethanolic extract. Vegetable and fruit ethanolic extracts that exhibited an antimutagenic effect (at the range 50-2000 microg/plate), in decreasing order, against NDMA and NPYR were as follows: licorice > kiwi > carrot and licorice > broccoli > pineapple > kiwi, respectively. Decreasing orders against NDBA and NPIP were, respectively, kiwi > onion > licorice = garlic > green pepper > carrot and licorice > garlic > pineapple > carrot.     

Determination of flunixin in edible bovine tissues using liquid chromatography coupled with tandem mass spectrometry

An accurate, reliable, and reproducible assay was developed and validated to determine flunixin in bovine liver, kidney, muscle, and fat. The overall recovery and percent coefficient of variation (%CV) of twenty-eight determinations in each tissue for flunixin free acid were 85.9% (5.9% CV) for liver, 94.6% (9.9% CV) for kidney, 87.4% (4.7% CV) for muscle, and 87.6% (4.4% CV) for fat. The theoretical limit of detection was 0.1 microg/kg (ppb, ng/g) for liver and kidney, and 0.2 ppb for muscle and fat. The theoretical limit of quantitation was 0.3, 0.2, 0.6, and 0.4 ppb for liver, kidney, muscle, and fat, respectively. The validated lower limit of quantitation was 1 ppb for edible tissues with the upper limit of 400 ppb for liver and kidney, 100 ppb for fat, and 40 ppb for muscle. Accuracy, precision, linearity, specificity, ruggedness, and storage stability were demonstrated. Briefly, the method involves an initial acid hydrolysis, followed by pH adjustment ( approximately 9.5) and partitioning with ethyl acetate. A portion of the ethyl acetate extract was purified by solid-phase extraction using a strong cation exchange cartridge. The eluate was then evaporated to dryness, reconstituted, and analyzed using LC/MS/MS. The validated method is sensitive and specific for flunixin in edible bovine tissue.     

Tissue fluoroacetate residues in prairie dogs dosed with low-level sodium monofluoroacetate

A total of 83 black-tailed prairie dogs (Cynomys ludovicianus) from South Dakota were subjected to low-level treatment with sodium monofluoroacetate (Compound 1080) in the laboratory (0.01-0.30 mg 1080/kg). The acute oral median lethal dose (LD50) of 1080 administered by oral gavage was established at 0.173 mg/kg. To assay fluoroacetate residues, 8 kinds of tissue from each of 10 prairie dogs dead of low-level 1080 poisoning were analyzed by capillary gas chromatography-mass spectrometry. Of the total of 79 tissues analyzed, 73 contained less than 100 ppb fluoroacetate, and 67 contained less than 50 ppb fluoroacetate. To test the effect of secondary poisoning on non-target species, 8 European ferrets (Mustela furo) were fed ground whole carcasses of prairie dogs dead of low-level 1080 poisoning, with no observable ill effects on the ferrets.     

Analysis of zearalenone and alpha-zearalenol in urine of ruminants using gas chromatography-tandem mass spectrometry

The present paper describes a sensitive procedure for quantitative analysis of the Fusarium mycotoxins zearalenone and alpha-zearalenol in urine of ruminants. Extraction is done with an octadecyl (C18) column and cleanup with a silica column providing a preparation that is analyzed by gas chromatography-tandem mass spectrometry (GC-MS/MS). The trimethylsilyl ether derivatives of zearalenone and alpha-zearalenol yield molecular ions with m/z 462 and 536, respectively. These ions are selected in the first mass analyzer and then fragmented in a collision cell to give characteristic daughter ions (m/z 151, 333, 318, and 446). The method is known as multiple reaction monitoring (MRM). Elimination of chemical background noise by selecting proper fragment ions produces chromatograms in which identification and quantitation in a biological matrix is possible. The method was tested with sheep urine from an experimental feeding trial and was used to confirm natural mycotoxicosis of cows affected with zearalenone. Zearalenone (1 ppb) and alpha-zearalenol (14 ppb) were found in 2 different cow urine samples. The detection limit for both zearalenone and zearalenol is 1 ppb (1 ng/mL) in urine and is linear between 1 and 20 ppb for the former and 1 and 10 ppb for the latter.     

Method for determination of pentobarbital in dry dog food by gas chromatography/mass spectrometry

A procedure has been developed and validated for measuring the concentration of pentobarbital residues in dry, extruded animal feed in the range of 3-200 ng/g (ppb) with an estimated limit of quantitation of 2 ppb. The method was developed for surveillance purposes: to measure the concentration of euthanizing agent which might be present in feeds incorporating rendered products which themselves might include some fraction of euthanized animals. A previously published qualitative procedure was modified by adding isotopically labelled pentobarbital as an internal standard. Dry feed was ground and extracted with methanol. The extract was loaded on a mixed-mode (C-18, anion exchange) solid-phase extraction cartridge designed for barbiturate residues. Pentobarbital was eluted and derivatized for gas chromatography/mass spectrometry in positive ion chemical ionization mode. Quantitation was based on the ratio of dimethyl-pentobarbital MH+ (m/z 255) vs dimethyl-pentobarbital-d(5) (m/z 260) in standards and extracts. Accuracy ranged from 112% at 3 ppb to 96% at 200 ppb, with relative standard deviations ranging from 4% at 3 ppb to 2% at 200 ppb.     

Determination and confirmation of 5-hydroxyflunixin in raw bovine milk using liquid chromatography tandem mass spectrometry

A method was developed and validated to determine 5-hydroxyflunixin in raw bovine milk using liquid chromatography tandem mass spectrometry (LC/MS/MS). The mean recovery and percentage coefficient of variation (%CV) of 35 determinations for 5-hydroxyflunixin was 101% (5% CV). The theoretical limit of detection was 0.2 ppb with a validated lower limit of quantitation of 1 ppb and an upper limit of 150 ppb. Accuracy, precision, linearity, specificity, ruggedness, and storage stability were demonstrated. A LC/MS/MS confirmatory method using the extraction steps of the determinative method was developed and validated for 5-hydroxyflunixin in milk from cattle. Briefly, the determinative and confirmatory methods were based on an initial solvent (acetone/ethyl acetate) precipitation/extraction of acidified whole milk. The solvent precipitation/extraction effectively removed incurred ((14)C) residues from milk samples. The organic extract was then purified by solid phase extraction (SPE) using a strong cation exchange cartridge (sulfonic acid). The final SPE-purified sample was analyzed using LC/MS/MS. The methods are rapid, sensitive, and selective and provide for the determination and confirmation of 5-hydroxyflunixin at the 1 and 2 ppb levels, respectively.     

Gas-liquid chromatographic determination of 3,6-dichloropicolinic acid in soils.

A gas-liquid chromatographic (GLC) method has been developed for the determination of the experimental herbicide 3,6-dichloropicolinic acid (Dowco 290) in soils. The method involves extraction of 1 g soil samples with 1N NaCl at approximately pH 7, methylation with diazomethane utilizing a microgenerator, and detection by electron capture GLC. Interferences are small, so that a cleanup step is not necessary even at the 6 ppb level. The procedure is rapid, requiring only 45 min/sample. Recoveries range from 84 to 94% at the 6-1000 ppb level with a minimum detectable limit of 6 ppb. Standard deviations for the percentage recovery values vary from 10.9 to 2.3 for the tested range of 6.7-670 ppb, respectively.     

Determination of Mirex in human blood serum containing polychlorinated biphenyls by using packed column gas chromatography.

An analytical method has been developed that uses electron capture/gas-liquid chromatography to determine Mirex in serum containing polychlorinated biphenyls (PCBs) (Aroclor 1260). With this method, 0.2 ppb Mirex can be determined in 4 mL serum that also contains 10 ppb PCBs. The method provides approximately 70% recovery of Mirex at 1.0 and 3.5 ppb. The coefficients of variation are 4.5 and 4.6% at 1.0 and 3.5 ppb, respectively. In a cooperative study with the Ohio Department of Health, the Centers for Disease Control used this method to determine the extent of exposure of Salem, OH, residents to Mirex. Confirmation of Mirex was obtained by using high resolution gas chromatography and high resolution mass spectrometry.     

Development of multiclass methods for drug residues in eggs: hydrophilic solid-phase extraction cleanup and liquid chromatography/tandem mass spectrometry analysis of tetracycline, fluoroquinolone, sulfonamide, and beta-lactam residues

A method was developed for detection of a variety of polar drug residues in eggs via liquid chromatography/tandem mass spectrometry (LC/MS/MS) with electrospray ionization (ESI). A total of twenty-nine target analytes from four drug classes-sulfonamides, tetracyclines, fluoroquinolones, and beta-lactams-were extracted from eggs using a hydrophilic-lipophilic balance polymer solid-phase extraction (SPE) cartridge. The extraction technique was developed for use at a target concentration of 100 ng/mL (ppb), and it was applied to eggs containing incurred residues from dosed laying hens. The ESI source was tuned using a single, generic set of tuning parameters, and analytes were separated with a phenyl-bonded silica cartridge column using an LC gradient. In a related study, residues of beta-lactam drugs were not found by LC/MS/MS in eggs from hens dosed orally with beta-lactam drugs. LC/MS/MS performance was evaluated on two generations of ion trap mass spectrometers, and key operational parameters were identified for each instrument. The ion trap acquisition methods could be set up for screening (a single product ion) or confirmation (multiple product ions). The lower limit of detection for screening purposes was 10-50 ppb (sulfonamides), 10-20 ppb (fluoroquinolones), and 10-50 ppb (tetracyclines), depending on the drug, instrument, and acquisition method. Development of this method demonstrates the feasibility of generic SPE, LC, and MS conditions for multiclass LC/MS residue screening.     

Sulfamethazine (sulfadimidine) residues in Canadian consumer milk

A survey on the presence of sulfamethazine (sulfadimidine) residues in consumer milk has been conducted in 10 cities across Canada. In each city, homogenized milk was purchased at 3 different retail outlets, each supplied by different processing plants. A total of 30 samples was analyzed by a liquid chromatographic method. The limit of quantitation was 5 ppb. In addition to automatic integration, visual inspection of the chromatograms was required to distinguish between low concentrations of sulfamethazine and 2 unknown interfering peaks. Two samples, from different cities, contained 11.4 and 5.24 ppb of the drug. Drug identity was confirmed by mass spectrometry. All other samples appeared to be free of the drug.     

Determination of bisphenol A in food-simulating liquids using LCED with a chemically modified electrode

Liquid chromatography with electrochemical detector (LC-ED), using a chemically modified electrode coated with a metalloporphyrin film, is reported for determination of bisphenol A (BPA) migration from polycarbonate baby bottles. The extraction process of the samples was performed according to regulations of the Southern Common Market (MERCOSUR), where certain food-simulating liquids [(A) distilled water, (B) acetic acid 3% V/V in distilled water, and (C) ethanol 15% V/V in distilled water] are defined along with controlled time and temperature conditions. The baseline obtained using the naked electrode showed a considerable drift which increased the detection limit. This effect was suppressed with the chemically modified electrode. A linear range up to 450 ppb along with a detection limit of 20 ppb for the amperometric detection technique was observed. The procedure described herein allowed lowering the detection limit of the method to 0.2 ppb. The value found for BPA in the food-simulating liquid is 1.2 ppb, which is below the tolerance limit for specific migration (4.8 ppm).     

Multiresidue pesticide analysis of agricultural commodities using acetonitrile salt-out extraction, dispersive solid-phase sample clean-up, and high-performance liquid chromatography-tandem mass spectrometry

A multiresidue method analyzing 209 pesticides in 24 agricultural commodities has been developed and validated using the original Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) procedure and high performance liquid chromatography-positive electrospray ionization-tandem mass spectrometry (LC-MS/MS) analysis. Using solvent-only calibration standards (SOCSs) and matrix-matched calibration standards (MMCSs), it was demonstrated that a minimal concentration of 5-10 μg/kg (part per billion, ppb) of analytes in matrix is required for the consistent identification of targeted pesticides with two MRM transitions. Method performance was validated by the precision and accuracy results obtained from fortification studies at 10, 25, 100, and 500 ppb and MMCSs. The method was demonstrated to achieve an average recovery of 100 ± 20% (n = 4) for >75% of evaluated pesticides at the low fortification level (10 ppb) and improved to >84% at the higher fortification concentrations in all 24 matrices. Matrix effects in LC-MS/MS analysis were studied by evaluating the slope ratios of calibration curves (1.0-100 ng/mL) obtained from the SOCSs and MMCSs. Principal component analysis (PCA) of LC-MS/MS and method validation data confirmed that each matrix exerts its specific effect during the sample preparation and LC-MS/MS analysis. The matrix effect is primarily dependent on the matrix type, pesticide type and concentration. Some caution is warranted when using matrix matched calibration curves for the quantitation of pesticides to alleviate concerns on matrix effects. The QuEChERS method with LC-MS/MS was used to identify and quantitate pesticides residues, with concentrations ranging from 2.5 to >1000 ppb in a variety of agricultural samples, demonstrating fitness for screening and surveillance applications.     

Effect of cooking on levels of ethylene dibromide residues in rice

Two studies were conducted to determine the effect that cooking has on the level of residues of ethylene dibromide (EDB) in rice. In the first study, 4 samples of long and medium grain polished white rice containing 113, 295, 956, and 1568 ppb EDB were cooked according to typical label directions. Three batches of cooked rice were prepared from each sample of polished rice and frozen until analysis; each batch was analyzed in duplicate. EDB levels in all cooked rice samples were less than 10 ppb. In the second study, conducted jointly by the Food and Drug Administration (FDA) and the Environmental Protection Agency (EPA), a sample of medium grain polished white rice containing about 1600 ppb EDB was cooked by each laboratory. Overall average EDB levels in rice analyzed immediately after cooking were 16 and 37 ppb for FDA and EPA, respectively. The corresponding frozen samples contained 8 and 39 ppb EDB. The 2 laboratories exchanged these frozen samples and reanalyzed them to check variability in the analytical procedure. FDA found 49 ppb EDB in the sample cooked by EPA and EPA found 8 ppb EDB in the sample cooked by FDA, thus indicating that analytical methodology was not a major source of variability. The range of EDB levels was therefore attributed to minor differences in the way the rice was cooked or handled immediately after cooking.