[Determination of Albendazole Metabolite in Livestock Products Using LC-MS/MS].

A method for determining albendazole metabolite (metabolite I) in livestock products using LC-MS/MS was proposed. Livestock samples were hydrolyzed with 6 mol/L HCl at 110℃ for an hour and defatted with ethyl acetate and n-hexane (1 : 1, v/v) mixture. Metabolite I was extracted with acetonitrile from the sample, and the extracts were salted out under basic conditions, allowing the acetonitrile layer to separate. The acetonitrile solution was cleaned up using a cartridge column packed with divinylbenzene-N-vinylpyrolidone copolymer bearing sulfo groups. The HPLC separation was conducted on an Inertsil ODS-4 column with a gradient formed from water containing 0.05% (v/v) formic acid and acetonitrile containing 0.05% (v/v) formic acid. To detect metabolite I, tandem mass spectrometry with positive ion electrospray ionization was used. Truenesses (n=5) of metabolite I from cattle meat, fat, liver, and milk spiked at the maximum residue limits or the 0.01 mg/kg were in the range from 83.6 to 97.9%, and the relative standard deviations were from 1.6 to 6.1%.

[Determination of Nonvolatile Amines in Foods by HPLC Following Fluorescamine Derivatization].

A method was developed for the determination of nonvolatile amines, such as histamine, tyramine, putrescine, and cadaverine, in foods. These nonvolatile amines were extracted from a sample with 5% trichloroacetic acid, and the extract was purified using an InertSep MC-1 cartridge column. The four amines were derivatized with fluorescamine, determined by HPLC with a fluorescence detector, and confirmed by LC-MS/MS. The average recoveries (n=5) and the relative standard deviations from 11 foods (pacific saury, dried mackerel, canned mackerel in brine, canned tuna in oil, fish sauce, surimi, rice-koji miso, soy sauce, gouda cheese, red wine, and beer) spiked at 100 mg/kg were 81-100% and 0.4-3.1%, respectively.

[Determination of Hexazinone and Its Major Metabolites in Livestock Products by LC-MS/MS].

A method for the determination of hexazinone and three metabolites (hexazinone metabolite B, hexazinone metabolite C, hexazinone metabolite F) in livestock products by LC-MS/MS was developed. Hexazinone and the three metabolites were extracted from a sample with acetonitrile in the presence of n-hexane, and lipid was removed by acetonitrile/n-hexane partition. The acetonitrile extract was cleaned up using a SAX/PSA cartridge column. Average recoveries (n=5) of hexazinone and the three metabolites from cattle meat, fat, liver and milk spiked at the maximum residue limits (MRLs) or at 0.0025 mg/kg ranged from 85.6 to 96.0%, and the relative standard deviations ranged from 0.8 to 4.9%.

Genetic engineering of novel flower colour by suppression of anthocyanin modification genes in gentian.

Ornamental gentian plants have vivid-blue flowers. The main factor contributing to the flower colour is the accumulation of a polyacylated delphinidin 'gentiodelphin' in their petals. Although in vitro studies proposed that acylation plays an important role in the stability and development of gentian blue colour, the in vivo stability of the polyacylated anthocyanin was not clearly demonstrated. Thus, to reveal the importance of anthocyanin modification, especially acylation, and to engineer new colours of gentian flowers, we used chimeric RNAi technology to produce transgenic gentian plants with downregulated anthocyanin 5,3'-aromatic acyltransferase (5/3'AT) and flavonoid 3',5'-hydroxylase (F3'5'H) activities, which are both essential enzymes for gentiodelphin biosynthesis. Two lines of flower colour-modified plants were obtained from fifteen transgenic gentian plants. Clone no. 1 exhibited a lilac flower colour and clone no. 15 exhibited pale-blue flowers. RNA gel blot analysis confirmed that both transgenic lines had markedly suppressed 5/3'AT transcripts, whereas clone no. 15 had fewer F3'5'H transcripts than clone no. 1 and untransformed control plants. HPLC analysis of anthocyanin compositions showed that downregulation of the 5/3'AT gene led to increased accumulation of non-acylated anthocyanins, as expected. These results demonstrated that genetic engineering to reduce the accumulation of polyacylated anthocyanins could cause modulations of flower colour.

Prevalence and risk factors for cornea guttata in the Reykjavik Eye Study.

PURPOSE: To establish the age- and gender-specific prevalence of cornea guttata (CG) in citizens of Reykjavik, Iceland, 55 years and older. DESIGN: Cross-sectional, random, population-based study. PARTICIPANTS: The 774 participants were those participating in the second examination of the Reykjavik Eye Study. At baseline, we had a response rate of 75.8%, and at the 5-year follow-up, 88% of the survivors participated. METHODS: We used slit-lamp and non-contact specular microscopy and endothelial specular photography as well as computer-assisted morphometry. We used a standardized grading system for CG. MAIN OUTCOME MEASURES: Diagnosis of primary central CG. RESULTS: The prevalence of CG is 11% for females and 7% for males both for right eyes and left eyes. Higher weight and higher body mass index are found to be associated with decreased risk of CG. Having smoked more than 20 pack-years increased the risk of CG more than 2-fold (P<0.02). CONCLUSIONS: Cornea guttata seem to be found more commonly in women than in men. Smoking for more than 20 pack-years increases the risk of developing CG more than 2-fold.

Parathyroid hormone-related protein (PTHrP) produced by dog lymphoma cells.

Parathyroid hormone-related protein (PTHrP) was investigated in a canine lymphoma case with hypercalcemia by means of immunoradiomentric assay (IRMA) and molecular analysis. The plasma calcium level of the patient dog was 13.7 mg/dl. The PTHrP concentration examined by IRMA was 6.1 pmol/L in the plasma sample from the dog, but it was undetectable (< 1.1 pmol/L) in plasma samples from 4 lymphoma cases without hypercalcemia or 5 normal dogs. The PTHrP concentration examined in the culture supernatant of the lymphoma cells from this case was 1.3 pmol/L, whereas those of the lymphoma cells from a lymphoma case without hypercalcemia was undetectable. PTHrP mRNA was clearly detected not only in the lymphoma cells from this dog with hypercalcemia but also in lymphoma cells from 4 lymphoma cases without hypercalcemia and 2 canine lymphoma cell lines.