Masakazu Horie

Determination of bisphenol A in canned vegetables and fruit by high performance liquid chromatography

A high performance liquid chromatograph y (HPLC) method was developed for the determination of bisphenol A (BPA) that had migrated into canned fruit and vegetables. BPA was extracted with acetonitrile from the solid portion of canned food, and with an OASIS HLB cartridge from the aqueous portion, respectively. Both extracts were cleaned up on a Florisil cartridge. The HPLC separation was carried out on a Wakosil II 3C18 RS column (4.6 x 150mm) with acetonitrile-water (40:60, v/v) as a mobile phase with a flow rate of 0.8 ml/min. BPA was detectable by UV detector at 228 nm and determined with the similarity of chromatographic peak spectrum by multiwavelength detector (similarity index was 0.99 or above). The quantification limits were 10 ng/g for the solid portion and 5 ng/ml for the aqueous portion, respectively. BPA was mainly detected in the solid portion of canned food and found at the maximum level of 11 μg per can. To verify migration into the solid portion of canned food, a partitioning experiment was carried out.

Simultaneous determination of five macrolide antibiotics in meat by high-performance liquid chromatography

A simple and rapid method using high-performance liquid chromatography (HPLC) for the simultaneous determination of five macrolides (josamycin, kitasamycin, mirosamicin, spiramycin and tylosin) in meat has been developed. The drugs were extracted with 0.3% metaphosphoric acid-methanol (7:3, v/v), and the extracts were cleaned up on a Bond Elut SCX (500 mg) cartridge. The HPLC separation was performed on a Puresil 5C18 column (150 x 4.6 mm I.D.) with a gradient system of 0.025 M phosphate buffer (pH 2.5)-acetonitrile as the mobile phase at a flow-rate of 1.0 ml/min. The drugs were detected at 232 mn for josamycin, kitasamycin, mirosamicin and spiramycin, and 287 mn for tylosin. The calibration graphs were rectilinear from 2.5 to 100 ng for each drug. The recoveries at the level of 1.0 microgram/g were 70.8-90.4%, and detection limits were 0.05 microgram/g for each drug.

High-performance liquid chromatographic determination of glycoalkaloids in potato products

Abstract A method for the determination of α-solanine and α-chaconine, two major potato glycoalkaloids, in commercial potato products by high-performance liquid chromatography (HPLC) was developed. The glycoalkaloids were extracted with methanol and then purified using Sep-Pak C18 or Sep-Pak NH2 cartridges. A Nucleosil 5-NH2 column was employed for HPLC with acetonitrile-20 mM potassium dihydrogenphosphate (75:25, v/v) as the mobile phase. The calibration graph was linear in the range 1–50, μ g/ml for both α-solanine and α-chaconine. The average recoveries were 82.4–92.6% for α-solanine and 86.5–97.4% for α-chaconine added to various commercial potato products at a level of 5 mg per 100 g. The proposed method is applicable to all commercially available potato products and potato starch.

Migration of 4-nonylphenol from polyvinyl chloride food packaging films into food simulants and foods

Migration of 4-nonylphenol (NP) from polyvinyl chloride (PVC) films for food packaging into food simulants and foods has been studied in domestic applications such as wrapping of food and reheating in a microwave oven. The migration of NP from the PVC films was determined by high-performance liquid chromatography with electrochemical coulometric-array detection (LC/ED). Twelve PVC films intended for commercial use and ten for domestic applications (total: 22 samples) were analysed. Some of the PVC films (two home-use and ten retail-use) contained NP at concentrations of between 500 and 3300 µg/g. Migration of NP from the films was influenced by the test conditions (n-heptane at 25°C for 60min, distilled water at 60°C for 30min and 4% acetic acid at 60°C for 30min). The amount of NP migrating from the PVC films into n-heptane (0.33-1.6 µg/cm2) was higher than the amount migrating into distilled water or 4% acetic acid (up to 9.7ng/cm2) for the 11 films in which NP was detected. Up to 0.23% of the NP migrated into distilled water and 4% acetic acid and up to 62.5% into n-heptane. In addition, we investigated NP migration into cooked rice samples wrapped in PVC film. Using spiked samples the method gave an average recovery of 83.7% (n = 5) with a standard deviation of 2.5%. Migration of NP ranged from not detectable (<1.0ng/g) to 410.0ng/g by reheating samples in a microwave oven for 1min and from not detectable to 76.5ng/g by keeping samples at room temperature for 30min.

Simultaneous determination of benofloxacin, danofloxacin, enrofloxacin and ofloxacin in chicken tissues by high-performance liquid chromatography

A simple, rapid and reliable high-performance liquid chromatographic (HPLC) method for the simultaneous determination of residual fluoroquinolones (benofloxacin, danofloxacin, enrofloxacin and ofloxacin) in chicken has been developed. The drugs were extracted with 0.2% metaphosphoric acid-acetonitrile (7:3, v/v), followed by a Bond Elut C18 clean-up procedure. The HPLC separation was carried out on a Wakosil II 5C18-HG column (150 x 4.6 mm I.D.) with 0.05 M phosphate buffer (pH 2.4)-acetonitrile (80:20, v/v) containing 2.5 mM 1-heptanesulfonic acid as the mobile phase. A fluorescence detector was used at an excitation wavelength of 295 nm and an emission wavelength of 455 nm. The calibration graphs were linear from 0.1 to 10 ng for danofloxacin and from 1 to 100 ng for benofloxacin, enrofloxacin and ofloxacin. The recoveries of the drugs from tissues fortified at a level of 0.2 microgram/g were 81.1-89.6%, and the detection limits were 0.01 microgram/g for ofloxacin, danofloxacin and enrofloxacin and 0.02 microgram/kg for benofloxacin. The time needed per sample was less than 60 min.

Multi-residue quantitation of aminoglycoside antibiotics in kidney and meat by liquid chromatography with tandem mass spectrometry

Quantitative methods using liquid chromatography coupled with tandem mass spectrometry were developed for seven kinds of aminoglycoside antibiotics in kidney and muscle tissues. Mass spectral acquisition was performed in the positive-ion mode by applying multiple reaction monitoring. Liquid chromatographic separation employed a ZIC®-HILIC column (SeQuant) for hydrophilic interaction chromatography. Extraction of the aminoglycosides was performed using liquid extraction with a phosphate buffer containing trichloroacetic acid, followed by a solid-phase clean-up procedure on a weak cation-exchange column with carboxypropyl (CBX) SPE cartridge (Mallinckrodt Baker). The limits of quantification were 25 ng g−1 for gentamicin, 50 ng g−1 for spectinomycin, dihydrostreptomycin, kanamycin and apramycin, and 100 ng g−1 for streptomycin and neomycin. These are well below the maximum residue limits set by the Codex Alimentarius Commission. The recoveries of all compounds from all tissues fortified at the level of quantification limits of 500 and 1000 ng g−1 were >70%, and the variability (relative standard deviation) was generally <12%.

Determination of trenbolone and zeranol in bovine muscle and liver by liquid chromatography–electrospray mass spectrometry

A sensitive and selective method using liquid chromatography (LC)-electrospray mass spectrometry for the determination of growth promoters, trenbolone and zeranol, in bovine muscle and liver has been developed. The LC separation was performed on a Zorbax XDB-C18 column (150x2.1 mm I.D.) using 0.005% acetic acid-acetonitrile (60:40, v/v) as the mobile phase at a flow-rate of 0.2 ml/min. The positive ionization produced typical (M+H)+ molecular ions of alpha-trenbolone and beta-trenbolone. On the other hand, the negative mode produced (M-H)- ion of zeranol. The calibration graphs for alpha-trenbolone, beta-trenbolone and zeranol were rectilinear from 2.5 pg to 1.0 ng with selected ion monitoring. The drugs were extracted with 0.2% metaphosphoric acid-acetonitrile (6:4, v/v), and the extracts were cleaned up on a OASIS HLB (60 mg) cartridge. The recoveries of the hormones from bovine muscle fortified at 2 ng/g were 82.3-85.1%, and detection limits were 0.5 ng/g for each drug.

Simultaneous determination of nalidixic acid, oxolinic acid and piromidic acid in fish by high-performance liquid chromatography with fluorescence and uv detection

A simple and rapid method for the simultaneous determination of nalidixic acid (NA), oxolinic acid (OXA) and piromidic acid (PMA) in cultured fish has been developed by high-performance liquid chromatography (HPLC). The drugs were extracted with 0.1% metaphosphoric acid-methanol (6:4), followed by a Sep-Pak C18 clean-up procedure. The HPLC separation was carried out on a Kaseisorb LC ODS 300-5 column (25 cm x 4.6 mm I.D.) using 5 mM phosphate buffer-acetonitrile (6:4) as a mobile phase. A fluorescence detector was used for NA and OXA at the excitation wavelength of 325 nm and the emission wavelength of 365 nm and an ultraviolet detector at 280 nm for PMA. The calibration graphs were rectilinear from 1 to 10 ng for OXA, from 2 to 20 ng for NA and PMA. The recoveries of NA, OXA and PMA added to fish were 81.5-85.3, 83.7-88.7 and 80.9-84.9%, respectively, with high accuracy. The limits of detection were 0.01 micrograms/g for each drug.

Simultaneous determination of residual synthetic antibacterials in fish by high-performance liquid chromatography

A simple and rapid high-performance liquid chromatographic (HPLC) method for the simultaneous determination of sulphamonomethoxine (SMMX), sulphadimethoxine (SDMX), sulphisozole (SIZ), nalidixic acid (NA), oxolinic acid (OXA), piromidic acid (PMA), furazolidone (FZ) and sodium nifurstyrenate (NFSA) in cultured fish was developed. The drugs were extracted with 0.2% metaphosphoric acid-methanol (6:4), followed by a Bond Elut C18 clean-up procedure. The HPLC separation was carried out on an Inertsil ODS column (150 x 4.6 mm I.D.) using 5 mM aqueous oxalic acid-acetonitrile (55:45) as the mobile phase with detection at 265 nm (0.04 a.u.f.s.). The calibration graphs were rectilinear from 1 to 20 ng for OXA, from 2 to 50 ng for SMMX, SDMX, SIZ, NA, PMA and FZ and from 5 to 100 ng for NFSA. The recoveries of each drug added to fish were 65.0-89.5%. The detection limits were 0.02 micrograms/g for OXA, 0.05 micrograms/g for SMMX, SDMX, SIZ, NA, PMA and FZ and 0.1 micrograms/g for NFSA.

Inhibition of lipopolysaccharide-induced pro-inflammatory cytokine expression via suppression of nuclear factor-κB activation by Mallotus japonicus phloroglucinol derivatives

An aqueous acetone extract obtained from the pericarps of Mallotus japonicus (MJE) was observed to inhibit pro-inflammatory cytokine (tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) production in a lipopolysaccharide (LPS)-activated murine macrophage-like cell line, RAW 264.7, or human blood monocytes. Several phloroglucinol derivatives were isolated from the pericarps as active compounds. Among these compounds, isomallotochromanol and isomallotochromene were the most potent in inhibiting cytokine production. MJE and the phloroglucinol derivatives significantly reduced these cytokine mRNA expressions. Gel shift analysis revealed that stimulation of macrophages with LPS caused an increase in the DNA binding activity of nuclear factor-kappaB (NF-kappaB), which was inhibited by isomallotochromanol and isomallotochromene. Western blot analysis showed that LPS reduced the IkappaB-alpha level in macrophages, while 10 microM isomallotochromanol and 10 microM isomallotochromene attenuated the LPS-induced decrease in IkappaB-alpha protein. We conclude that these phloroglucinol derivatives inhibit pro-inflammatory cytokine production and mRNA expression via suppression of NF-kappaB activation in activated macrophages.