Nitrogen deposition differentially regulates the sensitivity of gross primary productivity to extreme drought versus wetness.

Global hydroclimatic variability is increasing with more frequent extreme dry and wet years, severely destabilizing terrestrial ecosystem productivity. However, what regulates the consequence of precipitation extremes on productivity remains unclear. Based on a 9-year field manipulation experiment on the Qinghai-Tibetan Plateau, we found that the responses of gross primary productivity (GPP) to extreme drought and wetness were differentially regulated by nitrogen (N) deposition. Over increasing N deposition, extreme dry events reduced GPP more. Among the 12 biotic and abiotic factors examined, this was mostly explained by the increased plant canopy height and proportion of drought-sensitive species under N deposition, making photosynthesis more sensitive to hydraulic stress. While extreme wet events increased GPP, their effect did not shift over N deposition. These site observations were complemented by a global synthesis derived from the GOSIF GPP dataset, which showed that GPP sensitivity to extreme drought was larger in ecosystems with higher N deposition, but GPP sensitivity to extreme wetness did not change with N deposition. Our findings indicate that intensified hydroclimatic variability would lead to a greater loss of land carbon sinks in the context of increasing N deposition, due to that GPP losses during extreme dry years are more pronounced, yet without a synchronous increase in GPP gains during extreme wet years. The study implies that the conservation and management against climate extremes merit particular attention in ecosystems subject to N deposition.

Identification of immune related gene signature for predicting prognosis of cholangiocarcinoma patients.

Objective: To identify the gene subtypes related to immune cells of cholangiocarcinoma and construct an immune score model to predict the immunotherapy efficacy and prognosis for cholangiocarcinoma. Methods: Based on principal component analysis (PCA) algorithm, The Cancer Genome Atlas (TCGA)-cholangiocarcinoma, GSE107943 and E-MTAB-6389 datasets were combined as Joint data. Immune genes were downloaded from ImmPort. Univariate Cox survival analysis filtered prognostically associated immune genes, which would identify immune-related subtypes of cholangiocarcinoma. Least absolute shrinkage and selection operator (LASSO) further screened immune genes with prognosis values, and tumor immune score was calculated for patients with cholangiocarcinoma after the combination of the three datasets. Kaplan-Meier curve analysis determined the optimal cut-off value, which was applied for dividing cholangiocarcinoma patients into low and high immune score group. To explore the differences in tumor microenvironment and immunotherapy between immune cell-related subtypes and immune score groups of cholangiocarcinoma. Results: 34 prognostic immune genes and three immunocell-related subtypes with statistically significant prognosis (IC1, IC2 and IC3) were identified. Among them, IC1 and IC3 showed higher immune cell infiltration, and IC3 may be more suitable for immunotherapy and chemotherapy. 10 immune genes with prognostic significance were screened by LASSO regression analysis, and a tumor immune score model was constructed. Kaplan-Meier (KM) and receiver operating characteristic (ROC) analysis showed that RiskScore had excellent prognostic prediction ability. Immunohistochemical analysis showed that 6 gene (NLRX1, AKT1, CSRP1, LEP, MUC4 and SEMA4B) of 10 genes were abnormal expressions between cancer and paracancer tissue. Immune cells infiltration in high immune score group was generally increased, and it was more suitable for chemotherapy. In GSE112366-Crohn's disease dataset, 6 of 10 immune genes had expression differences between Crohn's disease and healthy control. The area under ROC obtained 0.671 based on 10-immune gene signature. Moreover, the model had a sound performance in Crohn's disease. Conclusion: The prediction of tumor immune score model in predicting immune microenvironment, immunotherapy and chemotherapy in patients with cholangiocarcinoma has shown its potential for indicating the effect of immunotherapy on patients with cholangiocarcinoma.

Warming reduces the production of a major annual forage crop on the Tibetan Plateau.

Climate warming has been proposed to increase primary production of natural grasslands in cold regions. However, how climate warming affects the production of artificial pastures in cold regions remains unknown. To address this question, we used open-top chambers to simulate warming in a major artificial pasture (forage oat) on the cold Tibetan Plateau for three consecutive years. Surprisingly, climate warming decreased aboveground and belowground biomass production by 23.1%-44.8% and 35.0%-46.5%, respectively, without a significant impact on their ratio. The adverse effects on biomass production could be attributed to the adverse effects of high-temperatures on leaf photosynthesis through increases in water vapor pressure deficit (by 0.05-0.10 kPa), damages to the leaf oxidant system, as indicated by a 46.6% increase in leaf malondialdehyde content, as well as reductions in growth duration (by 4.7-6.7 days). The adverse effects were also related to exacerbated phosphorus limitation, as indicated by decreases in soil available phosphorus and plant phosphorus concentrations by 31.9%-40.7% and 14.3%-49.4%, respectively, and increases in the plant nitrogen: phosphorus ratio by 19.2%-108.3%. The decrease in soil available phosphorus concentration could be attributed to reductions in soil phosphatase activities (by 9.6%-18.5%). The findings of this study suggest an urgent need to advance agronomic techniques and cultivate more resilient forage genotypes to meet the increasing demand of forage for feeding livestock and to reduce grazing damage to natural grasslands on the warming-sensitive Tibetan Plateau.

[Determination of chlormequat chloride residues in animal derived foods by high performance liquid chromatography-tandem mass spectrometry].

A method for the determination of chlormequat chloride (CCC) residues in animal derived foods by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was developed. The samples were extracted with acetonitrile containing 1% (v/v) acetic acid and defatted with n-hexane, followed by clean-up on a cationic solid phase extraction column. The analytes were separated on a Venusil MP C18(2) column (150 mm×2.1 mm, 3 µm) under a gradient elution program using acetonitrile and 0.1% (v/v) formic acid aqueous solution as the mobile phases. Then, the analytes were detected by tandem mass spectrometry using a positive electrospray ionization (ESI+) source in the multiple reaction monitoring (MRM) mode. Matrix-matched internal standard calibration curves were used for quantitative analysis. The calibration curves showed good linearity in the range of 0.200-500 µg/L for CCC, with correlation coefficients (r2) no less than 0.9993. The limit of quantification (LOQ) of the method was 0.500 µg/kg. The average recoveries of CCC in pork, beef, mutton, chicken, egg, pig kidney, beef liver, sheep kidney, chicken liver and milk matrices at spiked levels of 0.500-500 µg/kg were 93.4%-101%, and the relative standard deviations were 2.3%-8.0%. The method has less matrix interference, with high sensitivity, accuracy and reliability, and it is suitable for the quantitative detection of CCC residues in animal derived foods.

[Simultaneous determination of nine artificial sweeteners in food by solid-phase extraction coupled with high performance liquid chromatography-tandem mass spectrometry].

A simple and rapid method based on solid-phase extraction and high performance liquid chromatography-tandem mass spectrometry for the simultaneous determination of nine artificial sweeteners (acesulfame-K, saccharin sodium, cyclamate, sucralose, aspartame, alitame, neotame, dulcin, neohesperidin dihydrochalcone) in various foods was developed. The sweeteners in food samples were extracted with triethylamine buffer solution (pH 4.5) and cleaned using an SPE column equipped with hydrophilic and lipophilic packing material. The analytes were separated on a Phenomenex Knietex® F5 column (100 mm×2.1 mm, 2.6 µm) using 0.1% (v/v) formic acid-5 mmol/L ammonium formate/methanol as a mobile phase for gradient elution, and then determined by tandem mass spectrometry in positive and negative ESI modes under multiple reaction monitoring (MRM). The internal standard method was used to further suppress the matrix effect. The method proved to be very effective in the removal of matrix interferences. Calibration curves were linear within a studied range of concentrations (r2>0.999) for the nine artificial sweeteners. The limits of detection (LODs) and limits of quantification (LOQs) were within 2-30 µg/kg and 6-100 µg/kg, respectively. The recoveries for the nine investigated sweeteners were within 86.3%-106.3% at three spiked levels, with relative standard deviations (RSDs) between 1.2% and 5.9%. The developed method is rapid, efficient, accurate, and reliable; it can also be applied for the rapid determination of other artificial sweeteners in a complex food matrix.

Trace analysis of estrogens in milk samples by molecularly imprinted solid phase extraction with genistein as a dummy template molecule and high-performance liquid chromatography-tandem mass spectrometry.

Dummy molecularly imprinted polymer microspheres (DMIPMS) towards estrogens were synthesized by Pickering emulsion polymerization employing genistein (GEN) as a dummy template molecule. The FTIR analysis indicated the successful preparation of the imprinted polymers, and the characterization results of scanning electron microscopy and nitrogen adsorption desorption measurement indicated that the obtained DMIPMS are in possess of regular spherical shapes, porous structures and narrow diameter distribution, a BET surface area of 402.74 m2 g-1, a total pore volume of 0.568 cm3 g-1 and a pore diameter of 3.62 nm. The binding capacity and selectivity of DMIPMS were investigated in equilibrium binding experiments and chromatographic evaluation experiments through scatchard analysis and molecular imprinting factor (IF) analysis, respectively. The MIPs showed high binding capacity and excellent selectivity towards seven selected natural and synthetic estrogens, which are Estrone (E1), 17ß-estradiol (ßE2), estriol (E3), ethinylestradiol (EE2), dienestrol(DS), diethylstilbestrol (DES), and hexestrol (HEX). A method for selective determination of seven estrogens in milk samples via dummy molecularly imprinted solid phase extraction coupled with HPLC-MS/MS was developed, which showed good linearity from 2 to 500 µg L-1 with a correlation coefficient (R2) of more than 0.999. The detection limits were within the range of 0.10-0.35 µg L-1 and the recoveries of the seven estrogens at three spiking levels (10,100,250 µg L-1) ranged from 88.9% to 102.3% with relative standard deviation (RSD, n = 5) for intra-day and inter-day assays varied from 0.8% to 4.5%. The developed method is thus proven to be efficient and reliable for regular monitoring of trace estrogens in complex matrices such as milk samples.

Determination of ß-Agonist Residues in Animal-Derived Food by a Liquid Chromatography-Tandem Mass Spectrometric Method Combined with Molecularly Imprinted Stir Bar Sorptive Extraction.

A novel clenbuterol molecularly imprinted polymer (MIP)-coated stir bar was prepared and applied to the determination of six ß-agonists in animal-derived food. Characterization and various parameters affecting adsorption and desorption behaviours were investigated. The extraction capacities of clenbuterol, salbutamol, ractopamine, mabuterol, brombuterol, and terbutaline for MIP coating were 3.8, 2.9, 3.1, 3.5, 3.2, and 3.3 times higher, respectively, than those of the NIP coating, respectively. The method of MIP-coated SBSE coupled with HPLC-MS/MS was developed. The recoveries in pork and liver samples were 75.8-97.9% with RSD from 2.6 to 5.3%. Limits of detection (LODs) and limits of quantification (LOQs) were 0.05-0.15 µg/kg and 0.10-0.30 µg/kg, respectively. Good linearities were obtained for six ß-agonists with correlation coefficients (R2) higher than 0.994. These results indicated the superiority of the proposed method in the analysis of ß-agonists in a complex matrix.