Identification and quantification of phenolic acids and flavonol glycosides in Tunisian Morus species by HPLC-DAD and HPLC–MS

The total polyphenol and flavonoids in leaves of Morus alba var. alba, Morus alba var. rosa and Morus rubra were determined and identification of their components was carried out. The total content of phenolics varied between 345.20 and 631.53 mg gallic acid equivalents (GAE)/100 g dry weight (DW) basis. The total amount of flavonoids ranged between 193.87 and 398.33 mg rutin equivalents (RE)/100 g DW. Thirteen compounds were isolated by chromatography, and their structures determined to be mainly flavonol glycosides and phenolic acids. Three novel components were identified as kaempferol-7-O-glucoside, quercetin-3-O-β-glucoside-7-O-α-rhamnoside and quercetin-3-O-rhamnoside-7-O-glucoside, for the first time from mulberry leaves. Others known compounds were also identified.     

A multi-residue method for the determination of 90 pesticides in matrices with a high water content by LC-MS/MS without clean-up

A method using QuEChERS extraction and LC-MS/MS in electrospray positive ionisation mode was developed and validated for the analysis of 90 pesticides in a high water content matrix (tomato) in a single chromatographic run. To assess the intra-laboratory reproducibility of the method, validation was conducted on four different days by two different analysts. The validation data was treated using a spreadsheet developed in-house, which sets the most appropriate model for linear fit by determining whether the residuals of the calibration curves are homocedastic or heterocedastic. A statistical test for the significance of regression was also carried out. Calibration was always matrix-matched and the curves were obtained over the range 0.0075-0.10 or 0.020-0.125 mg kg(-1). Identification of analytes was based on retention times and MRM ratios. Recoveries were assessed at four different levels for each analyte and were between 73 and 106%, with relative standard deviations under reproducibility conditions of <20%. The measurement uncertainties of the method for each pesticide analysed were below 50%. Previous validation of the same method, applied to papaya samples and satisfactory results obtained in various proficiency tests with different high water content matrices, demonstrated the applicability of the method to these classes of commodities, without clean-up. The validated method will be applied routinely in the pesticide residues monitoring programme that constitutes the National Residue and Contaminant Control Plan of Brazil.     

Risk assessment of methyl-mercury intake through cephalopods consumption in Portugal

The intake of methyl-mercury (methyl-Hg) through the consumption of three common cephalopod species, cuttlefish (Sepia officinalis), squid (Loligo vulgaris) and octopus (Octopus vulgaris), in Portugal as well as the associated probability of exceeding the provisional tolerable weekly intake (PTWI) were estimated by combining methyl-Hg contamination levels in these three cephalopods with constructed consumption scenarios and with a hypothesised consumption distribution for the general Portuguese population. It was found that squid presents no serious health concern with respect to methyl-Hg, but cuttlefish and octopus consumption should not exceed two 150 g meals per week. Moreover, the methyl-Hg risk assessment for Portuguese consumers showed no risk concerning the observed cephalopods consumption levels. However, besides methyl-Hg, other toxic metals present in cephalopods, such as cadmium, may be a serious health concern and the methyl-Hg risk can be compounded by the risk of other foods containing significant methyl-Hg levels, especially long-lived sea predators. Accordingly, a cautionary note must be attached to advised maximum consumptions, which may be revised by future studies. Tail estimation (TE) estimator was more accurate for lower probabilities, rendering accurate risk estimations different from zero. However, for higher probabilities, the much simpler plug-in (PI) estimator could be applied. Additionally, limitations of a deterministic approach were identified.     

Endocrine modulation of the recipient environment affects development of bovine testis tissue ectopically grafted in mice

Testis tissue xenografting is a powerful approach for the study of testis development and spermatogenesis, and for fertility preservation in immature individuals. In bovine testis xenografts, maturation and spermatogenesis are inefficient when compared to other species. To evaluate if exogenous modulation of the endocrine milieu in recipient mice will affect spermatogenic efficiency in xenografts from newborn calves, recipient mice were treated with the GnRH antagonist acyline (5 mg/kg s.c. every 2 weeks) to reduce testosterone production in xenografts, or with 6-N-propyl-2-thiouracil (PTU, 0.1% in drinking water for 4 weeks), to induce transient hypothyroidism in recipient mice respectively. Both treatments altered developmental parameters of testis xenografts and reduced germ cell differentiation. While the effects of acyline treatment can be attributed to inhibition of GnRH and gonadotropin action, lower Sertoli cell numbers and decreased seminiferous tubule length observed after PTU treatment were opposite to effects reported previously in rats. Regardless of treatment, Sertoli cells underwent only partial maturation in xenografts as Müllerian inhibiting substance and androgen receptor expression were lower than in donor and adult tissue controls respectively. In conclusion, although treatments did not result in improvement of maturation of bovine testis xenografts, the current study demonstrates that exogenous modulation of the endocrine milieu to affect xenograft development in recipient mice provides an accessible model to study endocrine control of spermatogenesis in large donor species.     

FGF10 inhibits dominant follicle growth and estradiol secretion in vivo in cattle

Fibroblast growth factors (FGFs) are involved in paracrine control of follicle development. It was previously demonstrated that FGF10 decreases estradiol (E(2)) secretion in granulosa cell culture and that theca cell FGF10 mRNA expression is decreased in healthy follicles from abattoir ovaries. The main objectives of this study were to evaluate FGF10 and FGFR2b mRNA expression during follicular development in vivo, to evaluate the effect of FGF10 on follicle growth using Bos taurus taurus cows as a model, and to gain more insight into the mechanisms through which FGF10 inhibits steroidogenesis. Messenger RNA encoding both FGF10 and FGFR2b (main FGF10 receptor) was significantly more expressed in subordinate follicles (SFs) than in dominant follicles (DFs). The intrafollicular injection of FGF10 into the largest growing follicle at 7-8?mm in diameter interrupted the DF growth in a dose-dependent manner (11±0.4, 8.3±1 and 5.9±0.3?mm for 0, 0.1, and 1?μg/ml FGF10, respectively, at 72?h after treatment; P<0.05). In a third experiment, follicles were obtained 24?h after FGF10 (1?μg/ml) or PBS treatment through ovariectomy. In theca cells, FGF10 treatment did not affect mRNA encoding steroidogenic enzymes, LHCGR and IGFBPs, but significantly upregulated FGF10 mRNA expression. The expression of CYP19A1 mRNA in granulosa cells was downregulated by FGF10 treatment, which was accompanied by a 50-fold decrease in E(2) production, and decreased cyclin D2 mRNA. These results have shown that FGF10 and its receptor FGFR2b are more expressed in SFs and provide solid in vivo evidence that FGF10 acts as an important regulator of follicular growth in cattle.     

Comparing the growth of a molecular semiconductor on amorphous and semi-crystalline polycarbonate substrates

Polymeric substrates are of importance in plastic electronics. However, polymeric surfaces can exhibit different morphologies depending on whether they are amorphous or semi-crystalline. This work focuses on the impact of the surface structure of bisphenol A polycarbonate substrates on the nucleation and growth of a p-type semi-conductor, namely zinc phthalocyanine (ZnPc). ZnPc films were deposited under high vacuum at different substrate temperatures on oriented semi-crystalline as well as amorphous substrates of PC. The oriented substrates of PC were prepared by a method combining mechanical rubbing and solvent induced crystallization: the substrates show a periodic and regular alternation of oriented crystalline lamellae. Grazing incidence X-ray diffraction shows that the crystalline lamellae of PC have a preferential (a, c) contact plane. Moreover, the substrates show a bilayer structure made of a 60 nm-thick semi-crystalline overlayer atop an amorphous underlayer. UV–vis spectroscopy shows that the polymorphism of the ZnPc films is not modified by the surface structure of the PC substrate (amorphous versus semi-crystalline). However, the statistical analysis of domain size and density versus substrate temperature T_s evidences different apparent activation energies of the growth mechanism. High Resolution Transmission Electron Microscopy suggests that twinning along a (2 ?1 0) plane accounts for the bifurcations of the in-plane b-axis direction of the ZnPc nanocrystals. On oriented substrates of PC, such bifurcations are partly suppressed by the nanocorrugation of the surface, resulting in larger apparent domain sizes and unidirectional in-plane orientation.     

Reduced progesterone concentration during growth of the first follicular wave affects embryo quality but has no effect on embryo survival post transfer in lactating dairy cows

Fertility of lactating dairy cows is associated with reduced progesterone (P(4)) concentration compared with nonlactating animals. The objective of the current study was to determine whether P(4) during growth of the first follicular wave (FFW) affects embryo quality. Lactating Holstein cows at 33±3 days post partum were allocated to one of three treatments. Cows in the FFW and FFW with P(4) (FFWP) treatments started the superstimulation protocol on day 1 of the estrous cycle and second follicular wave (SFW) cows started the superstimulation protocol on estrous cycle day 7. Cows were superstimulated with 400 mg of NIH-FSH-P1 (FSH) given twice daily for 5 days, two prostaglandin F(2α) (PGF(2α)) injections given with the ninth and tenth injections of FSH, GNRH given 48 h after the first PGF(2α) injection, and timed insemination 12 and 24 h after the GNRH injection. Cows in the FFWP treatment received two intravaginal P(4) inserts during the superstimulation. Embryos were recovered 6.5 days after artificial insemination and excellent/good and fair embryos were frozen and transferred. Blood was sampled daily from estrous cycle day 0 until insemination from donor cows. During the superstimulation protocol, P(4) was (P<0.01) greatest for SFW cows followed by FFWP and FFW cows respectively. The percentage of embryos-oocytes from SFW and FFWP cows classified as excellent/good and fair embryos was (P=0.02) greater than those of FFW cows. Pregnancy per embryo transfer was not (P≥0.73) affected by embryo donor treatment. Reduced embryo quality of cows induced to ovulate the follicles from the first follicular wave is a consequence of reduced P(4) during follicle growth.