Detection of antibiotics in chicken eggs obtained from supermarkets in Ho Chi Minh City, Vietnam.

The residual levels of antibiotics in Vietnamese eggs were monitored from 2014 to 2015. A total of 111 egg packages, distributed by 11 different companies, were collected from supermarkets in Ho Chi Minh City and the levels of 28 antibiotics were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) screening method. Sixteen samples tested positive for antibiotics; a total of eight compounds (enrofloxacin, ciprofloxacin, norfloxacin, sulfadimethoxine, sulfamethazine, sulfamonomethoxine, tilmicosin and trimethoprim) were detected. Enrofloxacin was detected in eight samples, with two samples exhibiting concentrations exceeding 1,000 µg kg-1. Tilmicosin was detected in three samples at a range of 49-568 µg kg-1. We observed that two of the 11 companies frequently sold antibiotic-contaminated eggs (detection rates of 56 and 60%), suggesting that a number of companies do not regulate the use of antibiotics in egg-laying hens. Our findings indicate that livestock farmers require instruction regarding antibiotic use and that continual antibiotic monitoring is essential in Vietnam.

Determination of irradiation histories of raw beef livers using liquid chromatography-tandem mass spectrometry of 5,6-dihydrothymidine.

A method for detecting irradiation histories of raw beef livers was developed by measuring 5,6-dihydrothymidine (DHdThd) using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Liver DNA was extracted using phenol-chloroform extraction followed by precipitation in 50% ethanol. DNA was then enzymatically digested and nucleosides were purified using an OASIS MCX column. DHdThd and thymidine (dThd) contents of resulting test solutions were analyzed using LC-MS/MS. DHdThd was detected specifically after γ-irradiation. Concentration ratios of DHdThd to dThd in the test solutions increased dose-dependently after irradiation at 1.0-11.3kGy, which included the practical dose for sterilization of 2-7kGy. Dose-response curves from beef livers of individual animals almost overlapped. Thus, this method is a candidate for the detection of irradiation histories of foods from which DNA can be extracted.

Monitoring of Antibiotic Residues in Aquatic Products in Urban and Rural Areas of Vietnam.

Antibiotic residues in aquatic products in Vietnam were investigated. A total of 511 fish and shrimp samples were collected from markets in Ho Chi Minh City (HCMC), Thai Binh (TB), and Nha Trang (NT) from July 2013 to October 2015. The samples were extracted with 2% formic acid in acetonitrile and washed with dispersive C18 sorbent. Thirty-two antibiotics were analyzed by LC-MS/MS. Of the 362 samples from HCMC, antibiotic residues were found in 53 samples. Enrofloxacin was commonly detected, at a rate of 10.8%. In contrast, samples from TB and NT were less contaminated: only 1 of 118 analyzed samples showed residues in TB and only 1 of 31 showed residues in NT. These differences were attributed to the local manufacturing/distribution systems. To understand the current status of antibiotic use and prevent adverse effects that may be caused by their overuse, continual monitoring is required.

[Validation Study for Analytical Method of Diarrhetic Shellfish Poisons in 9 Kinds of Shellfish].

A method was developed for the simultaneous determination of okadaic acid, dinophysistoxin-1 and dinophysistoxin-2 in shellfish using ultra performance liquid chromatography with tandem mass spectrometry. Shellfish poisons in spiked samples were extracted with methanol and 90% methanol, and were hydrolyzed with 2.5 mol/L sodium hydroxide solution. Purification was done on an HLB solid-phase extraction column. This method was validated in accordance with the notification of Ministry of Health, Labour and Welfare of Japan. As a result of the validation study in nine kinds of shellfish, the trueness, repeatability and within-laboratory reproducibility were 79-101%, less than 12 and 16%, respectively. The trueness and precision met the target values of notification.

Rapid and easy multiresidue method for the analysis of antibiotics in meats by ultrahigh-performance liquid chromatography-tandem mass spectrometry.

This study involved the development of a multiresidue method for the rapid analysis of 43 antibiotics in meats using ultrahigh-performance liquid chromatography-tandem mass spectrometry. This method was performed using dispersive-solid phase extraction, which is able to analyze 20 samples within 2 h. All compounds were determined simultaneously on a C18 separation column with gradient elution. Validation of the analytical method was performed by carrying out linearity, limit of quantification (LOQ), accuracy, precision, and recovery tests in different meat products. The validation criteria were set according to AOAC International and Japanese validation guidelines. The linearity of each compound was almost the coefficient of determination (r(2)) > 0.98. The LOQs of all tested antibiotics were <10 µg/kg. The results verify that this method is capable of quantitative analysis of 36, 33, and 37 compounds in beef, pork, and chicken, respectively. This method can be used for rapid and easy multiresidue screening of antibiotics for three meats (pork, beef, and chicken).

Antibiotic residue monitoring results for pork, chicken, and beef samples in Vietnam in 2012-2013.

A monitoring plan of residual antibiotics in food of animal origin was conducted in Vietnam from 2012 to 2013. Meat samples were collected from slaughterhouses and retail stores in Ho Chi Minh City and Nha Trang. A total of 28 antibiotics were analyzed using a LC-MS/MS screening method. Sulfonamides, fluoroquinolones, and tilmicosin were detected in some of the samples. Sulfaclozine and fluoroquinolones were mainly detected in chicken samples, and sulfamethazine was mainly detected in pork samples. High levels of sulfonamide residues, ranging between 2500 and 2700 µg/kg sulfaclozine and between 1300 and 3600 µg/kg sulfamethazine, were present in two chicken and three pork samples, respectively. Tilmicosin was detected at ranges of 150-450 µg/kg in 10 chicken samples. Positive percentages were 17.3, 8.8, and 7.4% for chicken, pork, and beef, respectively, for an average of 11.9%. The results suggest an appropriate withdrawal period after drug administration had not been observed in some livestock.

[Simultaneous determination of veterinary drugs in livestock products using dispersive and cartridge column solid-phase extraction by LC-MS/MS].

A method using dispersive and cartridge column solid-phase extraction (SPE) was developed for the simultaneous determination of veterinary drugs in livestock products by LC-MS/MS. The samples were extracted with 85% acetonitrile and cleaned up using dispersive and cartridge column SPE. Recoveries of 70 analytes fortified at the levels of 0.002 µg/g and 0.01 µg/g in cattle, swine and chicken muscles were examined. Among 70 analytes examined, 64 in cattle, 58 in swine and 49 in chicken satisfied the required Japanese validation guideline. The limit of quantitation values of the 70 analytes were 0.001 µg/g or 0.005 µg/g in three kinds of livestock products. This method is suitable to analyze 40 analytes in three kinds of livestock products at levels under the MRL.

[Validation study on a rapid multi-residue method for determination of pesticide residues in vegetables and fruits by LC-MS/MS].

A validation study was conducted on a rapid multiresidue method for determination of pesticide residues in vegetables and fruits by LC-MS/MS. Pesticide residues in the vegetables or fruits were extracted with acetonitrile in a disposable tube using a homogenizer, followed by salting out with anhydrous magnesium sulfate and sodium chloride in the presence of citrate salts for buffering. The extract was purified with a double-layered cartridge column (graphite carbon black/primary secondary amine silica gel; GCB/PSA). For citrus fruits a purification step with a C18 column was added (this column was connected to the GCB/PSA column). After removal of the solvent, the extract was resolved in methanol/water and analyzed by means of LC-MS/MS. The method was validated according to the method validation guideline of the Ministry of Health, Labour and Welfare of Japan; recovery tests were performed on 8 kinds of vegetables and fruits [cabbage, cucumber, Japanese radish, onion, potato, spinach, Amanatsumikan (a citrus fruit) and apple] by fortification of 161 pesticide residues at the concentrations 0.01 and 0.05 µg/g (each concentration of pesticide residue was extracted from 2 samples on 5 separate days). The trueness of the method for 127 pesticides in all 8 commodities was 70-120% with satisfactory repeatability and within-run reproducibility. This method is concluded to be applicable for determination of pesticide residues in vegetables and fruits.

[Trial for estimating the concentration of pesticide residues in the ingredients of processed foods].

If pesticide residues are detected at over the uniform limit (0.01 ppm) in processed foods, the maximum residue limits of pesticides in each ingredient should be considered. For that purpose, analysis of pesticide residues in the ingredients would be required, but in many cases it is difficult to obtain ingredients from the same lot, or the processed foods may have been manufactured overseas. Thus, a means of estimating pesticide concentrations in ingredients before processing would be important. In this study, we estimated the concentration of paclobutrazol, which was detected in LM pickles (LMPs), that would have been present in Takana (LM; mustard leaf) before processing. The LMPs mainly consisted of LM and seasoning liquid (SL), and other ingredients (sesame seeds or red pepper) accounted for less than 0.2% (w/w) of the LMPs. The LM and SL were separated and their paclobutrazol concentrations were determined. The concentration of paclobutrazol in the SL was approximately 1/10 of that detected in LM. Loss of water from LM during processing was taken into account to calculate the content of paclobutrazol in LM before processing, based on the analytical result of processed LM.

[Study of multi-residue method for determining pesticide residues in processed foods manufactured from agricultural products by LC-MS/MS].

A rapid multi-residue method for determination of pesticide residues in processed foods manufactured from agricultural products was examined. Five mL water was added to 5 g sample in a polypropylene tube, and the tube was left to stand at room temperature for 30 min. Then, 20 mL acetonitrile was added to the sample. The mixture was homogenized in a high-speed homogenizer, followed by salting out with 1 g NaCl and 4 g anhydrous MgSO4. After centrifugation, the organic layer was purified on a graphitized carbon/PSA cartridge column. After removal of the solvent, the extract was resolved in methanol-water and analyzed with LC-MS/MS. The recoveries of 93 pesticides fortified into 5 kinds of processed foods [Chinese cabbage kimchi, marmalade, raisin, umeboshi (pickled plum) and worcester sauce] were examined at the concentrations of 0.02 and 0.1 µg/g (n=5). The recoveries of 61 pesticides in all foods were 70-120% with relative standard deviation below 20% at both concentrations. Seventy-four processed foods obtained from markets in Japan were examined with this method. Pesticide residues over the maximum residue limit (0.01 µg/g) were detected in 2 processed foods.

[Application of a rapid and simple multi-residue method for determination of pesticide residues in drinking water and beverages using liquid chromatography-tandem mass spectrometry].

A rapid and simple multi-residue method for determination of pesticides has been applied to drinking water and beverages. To a disposable polypropylene tube containing 10.0 g sample, 20 mL acetonitrile was added and the mixture was shaken vigorously for 1 min to extract pesticides. Then, 1 g sodium chloride and 4 g magnesium sulfate anhydrous were added, followed by vigorous shaking for 1 min and centrifugation to obtain the organic phase. The organic phase was processed with a graphite carbon black/PSA solid phase column. After concentration and reconstitution with 25% methanol containing aqueous solution, the test solution was analyzed with LC-MS/MS. Recovery tests of 91 pesticides fortified (0.02 µg/g) in 35 kinds of drinking water and beverages were conducted. The decline of recoveries in alcoholic beverages is considered to be due to the increase of organic phase volume owing to ethanol included in the alcoholic beverages. A simulation study was carried out with simulated alcoholic beverages, which consisted of 50% grape juice, with various amounts of ethanol and water, to examine pesticides recoveries and volume of the organic phase. The results suggested this method would be applicable both to alcoholic beverages containing less than 10% ethanol and to alcoholic beverages containing over 10% ethanol after dilution with water to below 10% ethanol prior to the addition of acetonitrile. A sample could be processed and analyzed by LC-MS/MS within 2 h. Thus, this method should be useful for monitoring and screening pesticide residues in drinking water and various beverages.

[Preparation of samples for proficiency testing of pesticide residue analysis in processed foods].

To conduct proficiency testing for the analysis of pesticide residues in processed foods, fortified samples of retort curry and pancake were examined. In the case of retort curry, heating and mixing were necessary at the time of preparation to provide a homogenous analytical sample. A mixture of 4 carbamates and 11 organophosphorus pesticides was spiked and 14 of them showed consistent results in the samples. In the case of pancake, 10 kinds of pesticides were added to the pastry. The prepared pastry was them cooked. The relative concentrations of most of the pesticides in the pancake were not affected and all the pesticides showed consistent results in the samples. These results showed that the two tested samples were suitable for proficiency testing.

[Multiresidue method for determination of pesticide residues in processed foods by GC/MS].

A rapid and simple multiresidue method has been established for determination of pesticide residues in processed foods by GC/MS. The pesticides were extracted with ethyl acetate in the presence of anhydrous MgSO(4) in a disposable tube, using a homogenizer. The extract was concentrated and reconstituted in hexane, followed by acetonitrile-hexane partition to remove lipids. The acetonitrile layer was cleaned up with a double-layered SPE cartridge column (graphite carbon black/PSA silica gel). After removal of the solvent, the residue was dissolved in acetone. The test solution was subjected to GC/MS in the EI mode. Recovery tests of 222 pesticides from five kinds of processed foods (dumpling, curry, French fries, fried chicken, fried fish) were performed at two different fortification levels of 0.02 and 0.1 microg/g, and 100 pesticides showed acceptable recovery (70-120%) with low relative standard deviation (

[Multiresidue method for determination of pesticide residues in processed foods by GC/MS/MS].

A rapid and simple multiresidue method has been established for determination of pesticide residues in processed foods by GC/MS/MS. The pesticides were extracted with ethyl acetate, and were cleaned up with a double-layered SPE cartridge column (graphite carbon black/PSA silica gel). Recovery tests of 258 pesticides from five kinds of processed foods (dumpling, curry, French fries, fried chicken, fried fish) were performed at two different fortification levels of 0.02 and 0.1 microg/g, and 184 pesticides showed acceptable recovery (70-120%) with relative standard deviation of

Simple and rapid analysis of aristolochic acid contained in crude drugs and Kampo formulations with solid-phase extraction and HPLC photodiode-array detection.

Simple and rapid analysis of aristolochic acid (AA) in crude drugs and Kampo extracts using a solid-phase extraction method and HPLC-PDA analysis was investigated. Extraction of AA from samples was accomplished by adding methanol containing 1% ammonia. The addition of ammonia ionized the AA of acidic substances so that they adhered to an acrylamide copolymer of a strong anion exchange resin (Sep-Pak QMA) coupled to diol silica easily. Furthermore, a mixture of acetonitrile-water-phosphoric acid (75:25:2, v/v) was effective in isolating AA from its carrier. Since almost all interfering peaks originating from contaminants in crude drugs and Kampo extract formulations could be removed, a satisfactory HPLC chromatogram of AA was obtained. A good result was also obtained when Aristolochiaceae and crude drugs containing AA were tested. Particularly in the case of the medicinal parts of Asarum, several interfering peaks and a ghost peak detected near the AA peak were eliminated. The AA contents of two Kampo extract formulations, tokishigyakukagoshuyushokyoto and ryutanshakanto, were calculated by HPLC analysis. The AA content (the sum of AA-I and AA-II) was 1.25-6.13 mg per daily dose. From an additional recovery experiment for Kampo formulations, high recovery rates of AA were obtained. Neither LC/MS nor special instrumentation was necessary. Our results suggest that this simple, quick, and sensitive analytical method to detect AA in crude drugs and Kampo extract formulations would be valuable in safety inspections of AA in crude drugs and their products.

[Determination of melamine in Chinese-made processed food].

A method using liquid chromatograph coupled with tandem mass spectrometer (LC/MS/MS) was developed for the determination of melamine in processed food. After extraction with 50% acetonitrile and clean-up with PSA and SCX, the quantification limit of melamine in processed food was 0.5 microg/g. The recoveries and relative standard deviations were 113 to 115% and less than 5%, respectively. Therefore, we considered that the developed method offers high precision and sensitivity for the determination of melamine in processed food. When the method was applied to six suspected products, 0.8 to 37 microg/g of melamine was detected from four milk-rich products.

[Determination of pesticides in Chinese dumplings using liquid chromatography-tandem mass spectrometry].

A rapid and easy multiresidue method for determination of pesticide residues in Chinese dumplings using liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been developed. Pesticide residues were extracted with ethyl acetate in the presence of anhydrous magnesium sulfate in a disposable tube using a homogenizer. The extract was concentrated and reconstituted in hexane, followed by acetonitrile-hexane partition to remove lipids. The acetonitrile layer was purified with a double-layered cartridge column (graphite carbon black/primary secondary amine silica gel). After removal of the solvent, the extract was resolved in methanol/water and analyzed with LC-MS/MS. Recovery tests of 99 pesticide residues from Chinese dumpling were performed at 20 and 100 ng/g, and 72 pesticides exhibited acceptable recoveries (70-120%) with low relative standard deviations (<20%) at both concentrations. The time for sample preparation with 12 samples to test solutions was approximately 6 hr. This method could be useful for determination of pesticide residues in the Chinese dumplings.

A rapid and easy multiresidue method for the determination of pesticide residues in vegetables, fruits, and cereals using liquid Chromatography/tandem mass spectrometry.

The applicability of a rapid and easy multiresidue method for determination of pesticide residues in agricultural products by using liquid chromatography/tandem mass spectrometry (LC/MS/MS) was examined. Pesticide residues were extracted with acetonitrile in a disposable tube using a homogenizer, followed by salting out with anhydrous magnesium sulfate and sodium chloride. The extract was purified with a double-layered cartridge column (graphite carbon black/primary-secondary amine silica gel). After removal of the solvent, the extract was resolved in methanol-water and analyzed with LC/MS/MS. Recovery tests of 99 pesticide residues from 7 agricultural products were performed at 20 and 100 ng/g. Throughout all of the agricultural products tested, 47 pesticides exhibited satisfactory recoveries (70-120%) and relative standard deviations (<20%) at both concentrations. The time for processing of 12 samples to test solutions was approximately 2-3 h. This method could be useful for determination of pesticide residues in agricultural products.

Rapid analysis of 56 pesticide residues in natural medicines by GC/MS with negative chemical ionization.

Many methods for determining pesticide residues in food have been reported. Although natural medicines should be confirmed to be as safe as food, few methods for determining pesticide residues in natural medicines have been reported. In this study, 56 pesticides were detected in natural medicines with a simple, rapid sample preparation method. This study indicates that the proposed method is useful for analyzing pesticides in natural medicines.

Simultaneous analysis of 260 pesticide residues in agricultural products by gas chromatography/triple quadrupole mass spectrometry.

A method for simultaneous analysis of about 260 pesticides by gas chromatography coupled to tandem mass spectrometry (GC/MS/MS) with a triple quadrupole analyzer (QqQ) has been studied. The pesticides were extracted with acetonitrile and cleaned up by a bilayer cartridge. A single injection method was developed for the monitoring of all of the targeted pesticides. Two MS/MS transitions were selected for each analyte using the intensity ratio obtained from them as a confirmatory parameter. By using matrix-matched standards, 260 pesticides could be determined in most matrixes with recoveries of 70-120% and a standard deviation of < or = 20 at 2 different fortification levels of 0.02 and 0.1 microg/g. The developed method was applied to the monitoring of 173 agricultural product samples from the local market. The sensitivities of this method were lower than with most of the selective GC detectors, such as flame photometric or single MS. The selectivity of QqQ gives a very clean chromatogram, making compound identification and confirmation easy. The quick and reliable monitoring was achieved by combination with rapid extraction and cleanup.