Solid-Phase Extraction Combined with High Performance Liquid Chromatography-Diode Array Detector for Rapid Determination of Estrogens in Milk

Purpose: To develop a simple but effective method for the quantitative analysis of estrogens in milk.Methods: Solid-phase extraction method was employed for the extraction of the estrogen from milk and high performance liquid chromatography-diode array detector (HPLC-DAD) was used for the determination of estrogen.Results: Optimal chromatographic conditions were achieved on an Eclipse XDB-C18 column at a flow rate of 0.8 mL/min at room temperature. The products were monitored using a DAD detector set at 280 nm. The retention time of the three main estrogens were 15 min with a gradient program of acetonitrile/water, from 0 to 3 min, holding 35 % acetonitrile; from 4 to 7 min, ramped to 55 % acetonitrile; from 8 to 20 min, maintained at 55 % acetonitrile. The results showed that the calibration curve had good linearity within the concentration range of 0.5 - 8.0 μg/mL with correlation coefficient (R2) in the range of 0.9929-0.9936. The limit of detection was in the range of 0.025 - 0.045 μg/mL while mean recovery of estrogen from milk samples varied from 92.6 to 104.1 %. Satisfactory precision was obtained both for intra-assay (RSD, 1.8 to 4.4 %) and inter-assay (RSD, 2.0 to 4.7 %).Conclusion: The proposed method is environmentally friendly, inexpensive and convenient, and should be helpful in analyzing estrogens in biological, environmental and food samples.Keywords: Solid-phase extraction, Milk, Estrogens, High performance liquid chromatography-diode array detecto

Biološka karakterizacija izolata golubljeg paramiksovirusa 1 i analiza njegove patogenosti u SPF pilića

Pigeon paramyxovirus type 1 (PPMV-1) is considered as an antigenic variant of Newcastle Disease leading to high mortality and significant economic losses to the poultry industry. However, the pathogenicity of PPMV-1 to chickens is still unclear. Herein, we reported the biological characterization of the isolated PPMV-1 from the diseased pigeons suspected to Newcastle Disease and studied its pathogenicity in SPF chickens. The phylogenetic tree and evolution distances revealed that the new isolate belonged to a VI.2.1.1.2.2 sub-genotype of NDV. Despite the cleavage motif of the F protein containing the 112RRQKR↓F117 sequence associated with virulent NDV strains, and the value of MDT and ICPI of the isolate showing mesogenic characteristics, the challenge trial showed that the isolate had weak pathogenicity to chickens while causing lesions in multiple tissues and organs in pigeons.Golublji paramiksovirus tipa 1 (PPMV-1) smatra se antigenskom varijantom njukaslske bolesti koja uzrokuje visoku smrtnost i znatne ekonomske gubitke u peradarskoj industriji. Patogenost virusa PPMV-1 u pilića međutim još uvijek nije jasna. U ovom je radu provedena biološka karakterizacija PPMV-1 izoliranog iz golubova za koje se sumnja da su oboljeli od njukaslske bolesti te je istražena njihova patogenost u SPF pilića. Filogenetsko stablo i evolucijske udaljenosti otkrili su da novi izolat pripada podgenotipu VI.2.1.1.2.2 virusa njukaslske bolesti (NDV). Unatoč motivu u mjestu cijepanja F-proteina koji sadržava sekvenciju 112RRQKR↓F117 povezanu s virulentnim sojevima NDV-a, vrijednosti MDT-a i ICPI-ja izolata pokazale su mezogene značajke. Ovo probno istraživanje je pokazalo da su izolati PPMV-1 slabo patogeni u pilića, dok u golubova uzrokuju lezije na više organa i tkiva

Engineering NAD+ availability for Escherichia coli whole-cell biocatalysis: A case study for dihydroxyacetone production

Background: Whole-cell redox biocatalysis has been intensively explored for the production of valuable compounds because excellent selectivity is routinely achieved. Although the cellular cofactor level, redox state and the corresponding enzymatic activity are expected to have major effects on the performance of the biocatalysts, our ability remains limited to predict the outcome upon variation of those factors as well as the relationship among them. Results: In order to investigate the effects of cofactor availability on whole-cell redox biocatalysis, we devised recombinant Escherichia coli strains for the production of dihydroxyacetone (DHA) catalyzed by the NAD + -dependent glycerol dehydrogenase (GldA). In this model system, a water-forming NAD + oxidase (NOX) and a NAD + transporter (NTT4) were also co-expressed for cofactor regeneration and extracellular NAD + uptake, respectively. We found that cellular cofactor level, NAD + /NADH ratio and NOX activity were not only strain-dependent, but also growth condition-dependent, leading to significant differences in specific DHA titer among different whole-cell biocatalysts. The host E. coli DH5α had the highest DHA specific titer of 0.81\ua0g/g DCW with the highest NAD + /NADH ratio of 6.7 and NOX activity of 3900 U. The biocatalyst had a higher activity when induced with IPTG at 37\ub0C for 8\ua0h compared with those at 30\ub0C for 8\ua0h and 18\ua0h. When cells were transformed with the ntt4 gene, feeding NAD + during the cell culture stage increased cellular NAD(H) level by 1.44 fold and DHA specific titer by 1.58 fold to 2.13\ua0g/g DCW . Supplementing NAD + during the biotransformation stage was also beneficial to cellular NAD(H) level and DHA production, and the highest DHA productivity reached 0.76\ua0g/g DCW /h. Cellular NAD(H) level, NAD + /NADH ratio, and NOX and GldA activity dropped over time during the biotransformation process.Conclusions: High NAD + /NADH ratio driving by NOX was very important for DHA production. Once cofactor was efficiently cycled, high cellular NAD(H) level was also beneficial for whole-cell redox biocatalysis. Our results indicated that NAD + transporter could be applied to manipulate redox cofactor level for biocatalysis. Moreover, we suggested that genetically designed redox transformation should be carefully profiled for further optimizing whole-cell biocatalysis. \ua9 2013 Zhou et al.; licensee BioMed Central Ltd

Establishment of an efficient plant regeneration culture protocol and achievement of successful genetic transformation in Jatropha curcas L.

An efficient and reproducible protocol is described for shoot-bud regeneration and Agrobacterium tumefaciens-mediated genetic transformation of J. curcas. Treating the explants with high concentrations (5–120 mg/L) of TDZ for short durations (5–80 min) before inoculation culture increased significantly the regeneration frequency and improved the quality of the regenerated buds. The highest shoot-buds induction rate (87.35%) was achieved when petiole explants were treated with 20 mg/L TDZ solution for 20 min and inoculated on hormone-free MS medium for 30 days. Regenerated shoots of 0.5 cm or a little longer were isolated and grafted to seedling stocks of the same species, and then the grafted plantlets were planted on half-strength MS medium containing 0.1 mg/L IBA and 2 mg/L sodium nitroprusside (SNP). This grafting strategy was found to be very effective, to obtain that healthy grafted plantlets ready for acclimatization within 20 days. By the above mentioned protocol and with general Agrobacterium – mediated genetic transformation methods only 65 days were needed to obtain intact transgenic plants

Progress and challenges in maternal health in western China:a Countdown to 2015 national case study

Background China is one of the few Countdown countries to have achieved Millennium Development Goal 5 (75% reduction in maternal mortality ratio between 1990 and 2015). We aimed to examine the health systems and contextual factors that might have contributed to the substantial decline in maternal mortality between 1997 and 2014. We chose to focus on western China because poverty, ethnic diversity, and geographical access represent particular challenges to ensuring universal access to maternal care in the region. Methods In this systematic assessment, we used data from national census reports, National Statistical Yearbooks, the National Maternal and Child Health Routine Reporting System, the China National Health Accounts report, and National Health Statistical Yearbooks to describe changes in policies, health financing, health workforce, health infrastructure, coverage of maternal care, and maternal mortality by region between 1997 and 2014. We used a multivariate linear regression model to examine which contextual and health systems factors contributed to the regional variation in maternal mortality ratio in the same period. Using data from a cross-sectional survey in 2011, we also examined equity in access to maternity care in 42 poor counties in western China. Findings Maternal mortality declined by 8·9% per year between 1997 and 2014 (geometric mean ratio for each year 0·91, 95% CI 0·91–0·92). After adjusting for GDP per capita, length of highways, female illiteracy, the number of licensed doctors per 1000 population, and the proportion of ethnic minorities, the maternal mortality ratio was 118% higher in the western region (2·18, 1·44–3·28) and 41% higher in the central region (1·41, 0·99–2·01) than in the eastern region. In the rural western region, the proportion of births in health facilities rose from 41·9% in 1997 to 98·4% in 2014. Underpinning such progress was the Government’s strong commitment to long-term strategies to ensure access to delivery care in health facilities—eg, professionalisation of maternity care in large hospitals, effective referral systems for women medically or socially at high risk, and financial subsidies for antenatal and delivery care. However, in the poor western counties, substantial disparity by education level of the mother existed in access to health facility births (44% of illiterate women vs 100% of those with college or higher education), antenatal care (17% vs 69%) had at least four visits), and caesarean section (8% vs 44%). Interpretation Despite remarkable progress in maternal survival in China, substantial disparities remain, especially for the poor, less educated, and ethnic minority groups in remote areas in western China. Whether China’s highly medicalised model of maternity care will be an answer for these populations is uncertain. A strategy modelled after China’s immunisation programme, whereby care is provided close to the women’s homes, might need to be explored, with township hospitals taking a more prominent role

Rapid determination of 103 common veterinary drug residues in milk and dairy products by ultra performance liquid chromatography tandem mass spectrometry

A multi-residue method has been developed for the identification and quantification of 103 common veterinary drug residues in milk and dairy Products. This method was based on QuEChERS with dispersive solid-phase where C18 sorbent and anhydrous sodium sulfate were used to sample purification. After evaporation and reconstitution, the samples were analyzed by ultra-performance liquid chromatography-tandem mass spectrometry. The mean recovery results were all higher than 60% except ampicillin, pipemidic acid, enoxacin, and estriol, and the relative standard deviation was <20.0%. The limit of quantification ranged between 0.1 and 5 μg/kg for milk and between 0.5 and 25 μg/kg for milk powder. It was successfully used to detect residues of veterinary drug in real samples. This study proposes a simple and fast analytical method for monitoring multi-class veterinary drug residues to ensure food safety