Two rapid methods of sample preparation and analysis of fatty foods (e.g., milk, eggs, and avocado) were evaluated and compared for 32 pesticide residues representing a wide range of physicochemical properties. One method, dubbed the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method for pesticide residue analysis, entailed extraction of 15 g sample with 15 mL acetonitrile (MeCN) containing 1% acetic acid followed by addition of 6 g anhydrous magnesium sulfate and 1.5 g sodium acetate. After centrifugation, 1 mL of the buffered MeCN extract underwent a cleanup step (in a technique known as dispersive solid-phase extraction) using 50 mg each of C18 and primary secondary amine sorbents plus 150 mg MgSO4. The second method incorporated a form of matrix solid-phase dispersion (MSPD), in which 0.5 g sample plus 2 g C18 and 2 g anhydrous sodium sulfate was mixed in a mortar and pestle and added above a 2 g Florisil column on a vacuum manifold. Then, 5 × 2 mL MeCN was used to elute the pesticide analytes from the sample into a collection tube, and the extract was concentrated to 0.5 mL by evaporation. Extracts in both methods were analyzed concurrently by gas chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. The recoveries of semi-polar and polar pesticides were typically 100% in both methods (except that basic pesticides, such as thiabendazole and imazalil, were not recovered in the MSPD method), but recovery of nonpolar pesticides decreased as fat content of the sample increased. This trend was more pronounced in the QuEChERS method, in which case the most lipophilic analyte tested, hexachlorobenzene, gave 27 ± 1% recovery (n = 6) in avocado (15% fat) with a <10 ng/g limit of quantitation.

The present study was conducted to develop an effective method for establishment of porcine parthenogenetic embryonic stem cells (ppESCs) from parthenogenetically activated oocyte-derived blastocysts. The addition of 10% fetal bovine serum (FBS) to the medium on the 3rd day of oocyte culturing improved the development of blastocysts, attachment of inner cell masses (ICMs) onto feeder cells, and formation of primitive ppESC colonies. ICM attachment was further enhanced by basic fibroblast growth factor, stem cell factor, and leukemia inhibitory factor. From these attached ICMs, seven ppESC lines were established. ppESC pluripotency was verified by strong enzymatic alkaline phosphatase activity and the expression of pluripotent markers OCT3/4, Nanog, and SSEA4. Moreover, the ppESCs were induced to form an embryoid body and teratoma. Differentiation into three germ layers (ectoderm, mesoderm, and endoderm) was confirmed by the expression of specific markers for the layers and histological analysis. In conclusion, data from the present study suggested that our modified culture conditions using FBS and cytokines are highly useful for improving the generation of pluripotent ppESCs.

Low levels of phthalates, including di(2-ethylhexyl) phthalate (DEHP), in raw bovine milk were determined using gas chromatography/time-of-flight mass spectrometry (GC/TOF-MS). A fast and convenient process of sample treatment combined with TOF-MS analysis (medium resolution >5000), yielded good recoveries (85-125%) and low limits of detection (<0.002 mg kg(-1)). The most commonly used phthalate, DEHP, was found in 15 out of 30 samples monitored in this study. DEHP concentrations in raw milk ranged from not detected to 0.154 mg kg(-1), and the mean concentration was 0.057 mg kg(-1). The dietary intake of DEHP was about 0.004 mg kg(-1) body weight day(-1) if a child (24 months, 13 kg body weight) drinks 1 L day(-1) of milk that contains the mean concentration of DEHP found in raw milk. The estimated dietary intake corresponded to 8% of the European Union tolerable daily intake (TDI) of 0.05 mg kg(-1) body weight day(-1). Dimethyl phthalate (DMP) and di-n-butyl phthalate (DBP) were found from two and 20 samples, respectively, at low levels. Diethyl phthalate (DEP), butylbenzyl phthalate (BBP), and di-n-octyl phthalate (DnOP) were not found in any of the samples.

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