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Abstract The exploration potential within deep-water petroliferous basins holds great promise for oil and gas resources. However, the dearth of geochemical and isotopic data poses a formidable challenge in comprehending the intricate hydrocarbon charging processes, thereby impeding the comprehensive understanding of hydrocarbon accumulation mechanisms and models. Consequently, the establishment of robust source–reservoir relationships in deep-water petroliferous basins represents a pivotal challenge that significantly influences the exploration strategies and the comprehension of hydrocarbon enrichment dynamics within such basins. In this study, we introduce a novel approach, termed the “source–reservoir dynamic evaluation method,” tailored to investigate reservoir accumulation models in deep-water petroliferous basins. This method uses basin simulation technology to recover the thermal evolution history and hydrocarbon generation and expulsion history of source rocks, and on this basis delimits the hydrocarbon kitchen range. At the same time, the maturity of source rocks corresponding to crude oil and natural gas in typical reservoirs is calculated. Then, when the thermal evolution degree of source rocks adjacent to the reservoir reaches this maturity, the corresponding geological period is the main charging period of hydrocarbon. As a typical deep-water petroliferous basin, the Santos Basin in Brazil has abundant oil and gas reservoirs under the thick salt rock, but there are still some fundamental problems such as unclear oil–gas accumulation process and model. Therefore, in this paper, the main charging periods of typical hydrocarbon reservoirs are determined based on the internal relationship between the thermal evolution history of the main source rocks and the maturity of crude oil and natural gas, and then the hydrocarbon accumulation process is analyzed and the dynamic accumulation model is established. Finally, the favorable prospecting direction is pointed out. The results show that the oil and gas in the Barra Velha Formation in the Santos basin are mainly derived from the Itapema Formation lacustrine shale source rock, and the source rock is mainly developed in the Eastern Sag of the Central Depression, and its main hydrocarbon generation period is from the deposition period of Florianopolis Formation to the deposition period of Santos Formation. The main hydrocarbon expulsion period was from the deposition period of the Santos Formation to the Early deposition of the Iguape Formation. The oil and gas in the Barra Velha Formation were mainly charged from the Late deposition period of the Santos Formation to the Early deposition period of the Iguape Formation. During this period, the hydrocarbon migrated vertically along the normal fault formed in the rift period to the trap of the adjacent inheritance structural highs and accumulated in the reservoir, which was dominated by the accumulation model of the “lower generation-upper reservoir-salt cap”. Since the Barra Velha Formation has the characteristics of near-source accumulation, based on the hydrocarbon expulsion center and hydrocarbon expulsion intensity of the source rock of the Itapema Formation, the distribution ranges of 85% and 50% Pre-salt accumulation probability in the Santos basin were calculated by using the quantitative analysis model of the hydrocarbon distribution threshold. It is suggested that the next oil and gas exploration should be carried out in the paleo-structural highs and slope of Class I favorable area (the hydrocarbon accumulation probability is more than 85%) and Class II favorable area (the hydrocarbon accumulation probability is 85–50%).

Abstract The Santos Basin, located in the southeastern waters of Brazil, is a passive continental margin basin with the most abundant deepwater petroleum resources in the world discovered to date. However, few studies have been conducted on the present geothermal fields of the Santos Basin, which severely restricts the oil and gas resource evaluation of the basin. This study first utilizes 35 temperature data from 16 post-salt drilling wells and 370 temperature data from 31 pre-salt drilling wells to calculate the post-salt and pre-salt geothermal gradients and terrestrial heat flows in the Santos Basin. Then, the basin simulation software BasinMod 1D is used to quantitatively evaluate the impacts of salt rock sedimentation on the present geothermal fields and the maturity of pre-salt hydrocarbon source rocks. The results demonstrate that the present post-salt geothermal gradient in the Santos Basin is 2.20–3.97 °C/100 m, with an average value of 2.99 °C /100 m, and the post-salt terrestrial heat flow is 54.00–97.32 mW/m2, with an average value of 73.36 mW/m2, while the present pre-salt geothermal gradient is 2.21–2.95 °C/100 m, with an average value of 2.53 °C/100 m, and the pre-salt terrestrial heat flow is 61.85–82.59 mW/m2, with an average value of 70.69 mW/m2. These values are characteristic of a low-temperature geothermal field in a zone with a stable structure. The sedimentation of the salt rock causes a decrease in the temperature of the pre-salt strata, which inhibits pre-salt hydrocarbon source rock maturity, with an inhibition rate of up to 1.32%. The inhibition degree decreases with increasing salt rock thickness. At the same time, the salt rock thickness is positively correlated with the present surface heat flow. The unique distribution of the salt rock and related salt structures lead to present terrestrial heat flow differences among different structural units in the basin. This study is of great significance for evaluating and exploring the pre-salt oil and gas resources in the Santos Basin.

Temperature variation affects the growth performance of fish species due to growing constraints and trade-off on physiological functions. Here we experimentally investigated several metabolic and physiological parameters in Sinilabeo rendahli, an endemic fish species found in the Yangtze River in China. After a 14-day acclimation period, we measured routine metabolic rate (RMR), individual metabolic rate (MO2), temperature quotient (Q10), loss of equilibrium (LOE), and critical oxygen tension (Pcrit) at three different temperatures (15, 20, and 25°C). Moreover, we sampled the muscle tissue from juvenile S. rendahli under experimental conditions after 28 days of acclimation and performed transcriptome-RNA sequencing (RNA-seq). The Pcrit of the fish at the above acclimation temperatures were determined to be 1.07, 1.28, and 1.33 mg·L−1, respectively, and corresponded with increasing acclimation temperatures (15–25°C). RMR was positively correlated with Pcrit (r = 0.4711, P = 0.0201), negatively correlated with LOE (r = −0.4284, P = 0.0367), and significantly positively correlated with MRcrit (r = 0.8797, P < 0.001) at temperatures ranging from 15 to 25°C. In addition, a total of 4,710 differentially expressed genes (DEGs) were identified. The results of DEG analysis and KEGG clustering analysis indicated that energy metabolism played a central role in thermal stress in S. rendahli for the major upregulated genes. This was followed by autophagy, mitophagy, cardiac muscle contraction, extracellular matrix (ECM)-receptor interaction, and protein digestion and absorption. This study is significant for understanding the adaptive response of S. rendahli to thermal stress. Even more importantly, this study demonstrates that S. rendahli is more suitable for cold-water life.

Shading is an important technique to protect tea plantations under abiotic stresses. In this study, we analyzed the effect of shading (SD60% shade vs. SD0% no-shade) on the physiological attributes and proteomic analysis of tea leaves in November and December during low temperatures. The results revealed that shading protected the tea plants, including their soil plant analysis development (SPAD), photochemical efficiency (Fv/Fm), and nitrogen content (N), in November and December. The proteomics analysis of tea leaves was determined using tandem mass tags (TMT) technology and a total of 7263 proteins were accumulated. Further, statistical analysis and the fold change of significant proteins (FC < 0.67 and FC > 1.5 p < 0.05) revealed 14 DAPs, 11 increased and 3 decreased, in November (nCK_vs_nSD60), 20 DAPs, 7 increased and 13 decreased, in December (dCK_vs_dSD60), and 12 DAPs, 3 increased and 9 decreased, in both November and December (nCK_vs_nSD60). These differentially accumulated proteins (DAPs) were dehydrins (DHNs), late-embryogenesis abundant (LEA), thaumatin-like proteins (TLPs), glutathione S-transferase (GSTs), gibberellin-regulated proteins (GAs), proline-rich proteins (PRPs), cold and drought proteins (CORA-like), and early light-induced protein 1, which were found in the cytoplasm, nucleus, chloroplast, extra cell, and plasma membrane, and functioned in catalytic, cellular, stimulus-response, and metabolic pathways. In conclusion, the proliferation of key proteins was triggered by translation and posttranslational modifications, which might sustain membrane permeability in tea cellular compartments and could be responsible for tea protection under shading during low temperatures. This study aimed to investigate the impact of the conventional breeding technique (shading) and modern molecular technologies (proteomics) on tea plants, for the development and protection of new tea cultivars.

Abstract The emergence of new highly pathogenic and drug-resistant influenza strains urges the development of novel therapeutics for influenza A virus (IAV). Here, we report the discovery of an anti-IAV microbial metabolite called APL-16-5 that was originally isolated from the plant endophytic fungus Aspergillus sp. CPCC 400735. APL-16-5 binds to both the E3 ligase TRIM25 and IAV polymerase subunit PA, leading to TRIM25 ubiquitination of PA and subsequent degradation of PA in the proteasome. This mode of action conforms to that of a proteolysis targeting chimera which employs the cellular ubiquitin-proteasome machinery to chemically induce the degradation of target proteins. Importantly, APL-16-5 potently inhibits IAV and protects mice from lethal IAV infection. Therefore, we have identified a natural microbial metabolite with potent in vivo anti-IAV activity and the potential of becoming a new IAV therapeutic. The antiviral mechanism of APL-16-5 opens the possibility of improving its anti-IAV potency and specificity by adjusting its affinity for TRIM25 and viral PA protein through medicinal chemistry.

The rhizoma of Polygonatum odoratum (PO) is used to treat yin injuries of the lung and stomach in traditional Chinese medicine. The chemical constituents of this herb are steroidal saponins, homoisoflavanones, and alkaloids. Xiangyuzhu (XPO) and Guanyuzhu (GPO) are available in the market as two specifications of the commodity. Nonetheless, systematic research on the identification and comparison of chemical constituents of these two commercial specifications is yet lacking. Herein, an integrated method combing ultra-high-performance liquid chromatography-quadruple time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) with ultra-high-performance liquid chromatography-charged aerosol detection (UHPLC-CAD) was employed for the comprehensively qualitative and quantitative analyses of PO. A total of 62 compounds were identified by UHPLC-Q-TOF/MS, among which 13 potential chemical markers were screened out to distinguish two commercial specifications. Subsequently, the absolute determination method for polygodoraside G, polygonatumoside F, and timosaponin H1 was established and validated by UHPLC-CAD. The contents of the three compounds were 13.33–236.24 μg/g, 50.55–545.04 μg/g, and 13.34–407.83 μg/g, respectively. Furthermore, the ratio of timosaponin H1/polygodoraside G could be applied to differentiate the two specifications. Samples with a ratio 5 are considered GPO. Therefore, the above results provide a valuable means for the quality control of PO.

Tea is a vital beverage crop all over the world, including in China. Low temperatures restrict its growth, development, and terrestrial distribution, and cold event variability worsens cold damage. However, the physiological and molecular mechanisms of Camellia sinensis under shade in winter remain unclear. In our study, tea leaves were utilized for physiological attributes and transcriptome analysis in November and December in three shading groups and no-shade control plants. When compared to the no-shade control plants, the shading group protected tea leaves from cold damage, increased photochemical efficiency (Fv/Fm) and soil plant analysis development (SPAD), and sustained chlorophyll a, chlorophyll b, chlorophyll, and carotenoid contents by physiological mean. Then, transcriptome analysis revealed 20,807 differentially expressed genes (DEGs) and transcription factors (TFs) in November and December. A comparative study of transcriptome resulted in 3,523 DEGs and many TFs under SD0% vs. SD30%, SD0% vs. SD60%, and SD0% vs. SD75% of shading in November and December. Statistically, 114 DEGs were downregulated and 72 were upregulated under SD0% vs. SD30%. SD0% vs. SD60% resulted in 154 DEGs, with 60 downregulated and 94 upregulated. Similarly, there were 505 DEGs of which 244 were downregulated and 263 were upregulated under SD0% vs. SD75% of shading throughout November. However, 279 DEGs were downregulated and 105 were upregulated under SD0% vs. SD30%. SD0% vs. SD60% resulted in 296 DEGs, with 172 downregulated and 124 upregulated. Finally, 2,173 DEGs were regulated in December, with 1,428 downregulated and 745 upregulated under SD0% vs. SD75%. These indicate that the number of downregulated DEGs in December was higher than the number of upregulated DEGs in November during low temperatures. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of differentially expressed genes were highly regulated in the photosynthesis, plant hormone signal transduction, and mitogen-activated protein kinase (MAPK) signaling pathways. However, qRT-PCR and RNA-seq relative expression of photosynthetic (DEGs) Lhcb2 in both November and December, plant hormone (DEGs) BRI1 and JAZ in November and IAA and ERF1 in December, and key DEGs of MAPK signal transduction FLS2, CHIB, and MPK4 in November and RBOH, MKK4_5, and MEKK1 in December in three shading groups and no-shade control plants responded to tea cold tolerance. The enhanced expression of light-harvesting photosystem I gene Lhca5, light-harvesting photosystem II gene Lhcb2, and mitogen-activated protein kinases MEKK1 and MPK4/6 enhance the cold-tolerance mechanism of C. sinensis. These comprehensive transcriptomic findings are significant for furthering our understanding of the genes and underlying regulatory mechanisms of shade-mediated low-temperature stress tolerance in horticultural crops.

Magnesium-based hydrogen storage materials are considered as one of the most promising candidates for solid state hydrogen storage due to their advantages of high hydrogen capacity, excellent reversibility and low cost. In this paper, Mg91.4Ni7Y1.6 and Mg92.8Ni2.4Y4.8 alloys were prepared by melting and ball milling. Their microstructures and phases were characterized by X-ray diffraction, scanning electron microscope and transmission electron microscope, and hydrogen absorbing and desorbing properties were tested by the high pressure gas adsorption apparatus and differential scanning calorimetry (DSC). In order to estimate the activation energy and growth mechanism of alloy hydride, the JMAK, Arrhenius and Kissinger methods were applied for calculation. The hydrogen absorption content of Mg92.8Ni2.4Y4.8 alloy reaches 3.84 wt.% within 5 min under 350 ℃, 3 MPa, and the maximum hydrogen capacity of the alloy is 4.89 wt.% in same condition. However, the hydrogen absorption of Mg91.4Ni7Y1.6 alloy reaches 5.78 wt.% within 5 min, and the maximum hydrogen absorption of the alloy is 6.44 wt.% at 350 ℃ and 3 MPa. The hydrogenation activation energy of Mg91.4Ni7Y1.6 alloy is 25.4 kJ/mol H2, and the enthalpy and entropy of hydrogen absorption are -60.6 kJ/mol H2 and 105.5 J/K/mol H2, separately. The alloy begins to dehydrogenate at 210 ℃, with the dehydrogenation activation energy of 87.7 kJ/mol H2. By altering the addition amount of Ni and Y elements, the 14H-LPSO phase with smaller size and ternary eutectic areas with high volume fraction are obtained, which provides more phase boundaries and catalysts with better dispersion, and there are a lot of fine particles in the alloy, these structures are beneficial to enhance the hydrogen storage performance of the alloys.

To investigate the effect of feeding different diets (non-artificial food and formulated diet and their mix) on specific dynamic action (SDA), digestive capacity and growth performance in carnivorous fish species, we measured the postprandial metabolic response and meal remaining in the stomach, specific growth rate (SGR) and intestinal microbial diversity of juvenile southern catfish (Silurus meridionalis) fed loach meat, formulated diet and their mix at 25 °C. The peak metabolic rate (PMR) and peak metabolic scope (PMS) of both the formulated diet and mixed diet group were higher than those of the loach meat group, and the SDA duration of both the formulated diet and mixed diet groups was shorter than that of the loach meat group. The percentage food remaining in the stomach was the lowest in the formulated diet group and the highest in the loach meat group at 16 and 24 h postfeeding, and the gastric evacuation rate was the highest in the formulated diet group and the lowest in the loach meat group. Both SGR and feeding rate (FR) were the highest in the formulated diet group and the lowest in the loach meat group. The bacterial community richness and relative abundance at the phylum and genus levels in the gut microbiota were higher in most samples of formulated diet group and lower in all samples of the loach meat group. These results indicated that southern catfish fed formulated diet had a higher metabolic response, faster digestion process and better growth performance than those fed loach meat. We suggest that these differences were related to the macronutrient composition (i.e., the relative content of protein, carbohydrate and lipid), type of food, as well as ingredients of formulated diets and gut microbiota.

Summary: Protein disulfide isomerase (PDI) plays a key role in maintaining cellular homeostasis by mediating protein folding via catalyzing disulfide bond formation, breakage, and rearrangement in the endoplasmic reticulum. Increasing evidence suggests that PDI can be a potential treatment target for several diseases. However, the function of PDI in the peripheral sensory nervous system is unclear. Here we report the expression and secretion of PDI from primary sensory neurons is upregulated in inflammatory and neuropathic pain models. Deletion of PDI in nociceptive DRG neurons results in a reduction in inflammatory and neuropathic heat hyperalgesia. We demonstrate that secreted PDI activates TRPV1 channels through oxidative modification of extracellular cysteines of the channel, indicating that PDI acts as an unconventional positive modulator of TRPV1. These findings suggest that PDI in primary sensory neurons plays an important role in development of heat hyperalgesia and can be a potential therapeutic target for chronic pain.

AIM: To observe changes in the content of matrix metalloproteinases (MMPs) in the corneal stroma after corneal cross-linking (CXL) in rabbits, and further explore the corneal pathophysiological process after CXL. METHODS: Forty-two rabbits (42 eyes) were randomly divided into seven groups. One group served as the control group, while the other six groups were treated with CXL. The concentrations of MMPs in corneal stroma were evaluated through parallel reaction monitoring at baseline and 3, 7, 15, 30, 90, and 180d after treatment. RESULTS: The levels of MMP-2 in the corneal stroma of rabbits were 0.76±0.07, 2.78±1.39, 4.12±0.69, 2.00±0.29, 2.00±0.30, 1.22±0.18, and 1.35±0.18 (10-9 mol/g) at baseline and 3, 7, 15, 30, 90, and 180d after treatment, respectively. The contents of tissue inhibitor of metalloproteinase-1 (TIMP-1) were 1.83±0.26, 7.94±0.58, 6.95±2.64, 3.81±0.48, 3.07±0.92, 1.72±0.19, and 1.69±0.74 (10-9 mol/g), respectively. The ratios of MMP-2/TIMP-1 were 0.42±0.33, 0.36±0.20, 0.62±0.10, 0.54±0.15, 0.68±0.13, 0.71±0.10, and 0.68±0.09, respectively. After CXL, the expression of MMP-2 and TIMP-1 in the rabbit corneal stroma was initially increased and subsequently decreased. The levels of MMP-2 remained higher than those recorded at baseline 180d after treatment, but it was not statistically significant. The levels of TIMP-1 returned to baseline levels at 90d after treatment. The ratio of MMP-2/TIMP-1 started to rise from 7d after CXL. It was significantly higher than that calculated at baseline 30-180d after CXL. The results for MMP-1, -3, -7, -9, -13, and TIMP-2 were negative. CONCLUSION: CXL can lead to changes in the content of MMP-2 and TIMP-1 in the rabbit corneal stroma. The ratio of MMP-2/TIMP-1 remains higher versus baseline, indicating that MMP-2 is involved in the corneal pathophysiological process after CXL.

Despite significant scientific advances toward the development of safe and effective radiation countermeasures, no drug has been approved for use in the clinic for prevention or treatment of radiation-induced acute gastrointestinal syndrome (AGS). Thus, there is an urgent need to develop potential drugs to accelerate the repair of injured intestinal tissue. In this study, we investigated that whether some fractions of Traditional Chinese Medicine (TCM) have the ability to regulate intestinal crypt cell proliferation and promotes crypt regeneration after radiation. By screening the different supplements from a TCM library, we found that an active fraction of the rhizomes of Trillium tschonoskii Maxim (TT), TT-2, strongly increased the colony-forming ability of irradiated rat intestinal epithelial cell line 6 (IEC-6) cells. TT-2 significantly promoted the proliferation and inhibited the apoptosis of irradiated IEC-6 cells. Furthermore, in a small intestinal organoid radiation model, TT-2 promoted irradiated intestinal organoid growth and increased Lgr5+ intestinal stem cell (ICS) numbers. More importantly, the oral administration of TT-2 remarkably enhanced intestinal crypt cell proliferation and promoted the repair of the intestinal epithelium of mice after abdominal irradiation (ABI). Mechanistically, TT-2 remarkably activated the expression of ICS-associated and proliferation-promoting genes and inhibited apoptosis-related gene expression. Our data indicate that active fraction of TT can be developed into a potential oral drug for improving the regeneration and repair of intestinal epithelia that have intestinal radiation damage.

Previous studies have suggested that Lycium barbarum (L. barbarum) has a radioprotective function, although more in-depth investigation is still required. We investigated the radioprotective efficacy of extract of the fruits of L. barbarum (LBE) and its radioprotective mechanisms. Mice were exposed to 8.5 Gy, 5.5 Gy, or 6.0 Gy total body irradiation (TBI), and the survival rate, lymphocyte percentage, amount of cytokines, and viability of the irradiated cells, as well as the gut microbiome and fecal metabolomics were studied. LBE enhanced the survival of the mice exposed to 8.5 Gy γ-ray TBI or 5.5 Gy X-ray TBI. After 6.0 Gy γ-ray TBI, LBE exhibited good immunomodulatory properties, mainly characterized by the accelerated recovery of lymphocyte percentages, and the enhanced expression of immune-related cytokines. LBE reconstituted the gut microbiota of irradiated mice, increased the relative abundance of potentially beneficial genera (e.g., Turicibacter, Akkermansia), and decreased the relative abundance of potentially harmful bacterial genera (e.g., Rikenellaceae_RC9_gut_group). Beneficial regulatory effects of LBE on the host metabolites were also noted, and the major upregulated metabolites induced by LBE, such as Tetrahydrofolic acid and N-ornithyl-L-taurine, were positively correlated with the immune factor interleukin (IL)-6. In vitro, LBE also increased the vitality of rat small intestinal epithelial cells (IEC-6) after 4.0 Gy γ-ray irradiation and promoted the growth of Akkermansia muciniphila. These results confirmed a radioprotective function of LBE and indicated that the radioprotective mechanism may be due to immunomodulation and the synergistically modulating effect on the gut microbiota and related metabolites.

Climate change and extreme weather affect tea growing. A competitive tea market needs quick, short-term solutions. This study evaluates the effects of various shade nets under mild and extreme cold stress on tea leaf physiology, photosynthetic alterations, antioxidant activities, and physiochemical characteristics. Tea plants were treated with SD0 (0% non-shading), SD1 (30% shading), SD2 (60% shading), and SD3 (75% shading). The 30%, 60%, and 75% shade nets shielded tea leaves from cold damage and reduced leaf injury during mild and extreme cold conditions compared with SD0% non-shading. Shading regulates photochemical capacity and efficiency and optimizes chlorophyll a and b, chlorophyll, and carotenoid contents. Moreover, carbon and nitrogen increased during mild cold and decreased in extreme cold conditions. Shading promoted antioxidant activity and physiochemical attributes. In fact, under 60% of shade, superoxide dismutase, peroxidase, catalase, and ω-3 alpha-linolenic acid were improved compared with SD0% non-shading during both mild and extreme cold conditions. From these findings, we hypothesized that the effect of different shades played an important role in the protection of tea leaves and alleviated the defense mechanism for “Zhong Cha 102” during exposure to a cold environment.

Anemarrhena asphodeloides is an herb widely used to treat symptoms associated with diabetes in traditional Chinese medicine. However, its key components and metabolites have low bioavailability and poor host absorption. To clarify the anti-diabetic mechanism of A. asphodeloides extract (AAE), we examined the anti-diabetic effects of AAE in rats with diabetes induced by a high-fat diet and streptozotocin. Faeces levels of the main components and metabolites of AAE were significantly higher than levels in plasma, which indicated that gut microbiota might play important roles in its anti-diabetic effect. Microbiological studies showed that unabsorbed components increased the diversity of the gut microbiota, enriched potentially beneficial bacteria, and suppressed potentially harmful bacteria. In vitro studies showed that AAE promoted the proliferation of Blautia coccoides, a bacterium with positive implication for diabetes, in a dose-dependent manner. AAE also promoted pancreatic cell regeneration and restored the function of pancreatic islet cells via peroxiredoxin 4 overexpression. Overall, these results suggest that AAE alleviates diabetes via modulating gut microbiota and protein expression.

Traditional diosgenin manufacturing process has led to serious environmental contamination and wastewater. Clean processes are needed that can alternate the diosgenin production. The β-glucosidase FBG1, cloned from Fusarium sp. CPCC 400709, can biotransform trillin and produce diosgenin. In this study, Pichia pastoris production of recombinant FBG1 was implemented to investigate various conventional methanol induction strategies, mainly including DO-stat (constant induction DO), μ-stat (constant exponential feeding rate) and m-stat (constant methanol concentration). The new co-stat strategy combining μ-stat and m-stat strategies was then developed for enhanced FBG1 production during fed-batch high-cell density fermentation on methanol. The fermentation process was characterized with respect to cell growth, methanol consumption, FBG1 production and methanol metabolism. It was found that large amounts of formaldehyde were released by the enhanced dissimilation pathway when the co-stat strategy was implemented, and therefore the energy generation was enhanced because of improved methanol metabolism. Using co-stat feeding, the highest volumetric activity reached ∼89 × 104 U/L, with the maximum specific activity of ∼90 × 102 U/g. After 108 h induction, the highest volumetric production reached ∼403 mg/L, which was ∼91, 154, and 183 mg/L higher than the maximal production obtained at m-stat, μ-stat, and DO-stat strategies, respectively. FBG1 is the first P. pastoris produced recombinant enzyme for diosgenin production through the biotransformation of trillin. Moreover, this newly developed co-stat induction strategy represents the highest expression of FBG1 in P. pastoris, and the strategy can be used to produce FBG1 from similar Pichia strains harboring Fbg1 gene, which lays solid foundation for clean and sustainable production of diosgenin. The current work provides unique information on cell growth, substrate metabolism and protein biosynthesis for enhanced β-glucosidase production using a P. pastoris strain under controlled fermentation conditions. This information may be applicable for expression of similar proteins from P. pastoris strains.

Steroidal saponins are widely used as starting precursors and medical intermediates for the semi-/total-synthesis of hundreds of steroidal drugs. One such steroidal saponin is diosgenin, which has attracted significant attention due to the huge market demand in the pharmaceutical industry. Due to water waste and severe environmental pollution, the traditional diosgenin production process based on direct acid hydrolysis is no longer used. In this study, to develop a submerged fermentation (SmF) medium for clean diosgenin production via efficient microbial biocatalysis, the Box–Behnken design (BBD) in combination with the Plackett–Burman design (PBD) was used to determine the medium compositions for Fusarium strains. Three components (wheat bran, phosphate, and Tween-80) were determined as significant factors by the PBD. Using the BBD, the three significant factors were further optimized, and the optimum values were determined for maximal diosgenin production. With 21.16 g/L of wheat bran, 9.60 g/L of phosphate, and 1.97 g/L of Tween-80, the diosgenin yield was 2.28%, i.e., 3.17 mg/L/h. The experimental values agreed with the predicted values, representing a significant increase in diosgenin production compared to its production using the basic SmF medium. For the first time, we reported the development of a new medium for Fusarium strains to produce diosgenin via microbial biocatalysis of the root of Dioscorea zingiberensis C. H. Wright (DZW). A simple-composition, low-cost, and high-efficiency medium was developed for the first time for the SmF of Fusarium strains. The medium is considered useful for large-scale SmF and may be applicable to other fungi. This study lays a solid foundation for diosgenin production in an acid-free and wastewater-free way. It may also provide fundamental support for producing other value-added products via microbial biocatalysis of low-value materials by endophytic fungi.

AIM: To report the clinical results of iontophoresis-assisted epithelium-on corneal crosslinking (I-CXL) using 0.1% riboflavin in distilled water for progressive keratoconus. METHODS: In this prospective clinical study, we examined 94 eyes of 75 patients with progressive keratoconus who were treated with I-CXL using 0.1% riboflavin in distilled water. Best correct visual acuity (BCVA), Scheimpflug tomography, corneal topography, anterior segment optical coherence tomography, intraocular pressure, and endothelial cell density were evaluated at baseline and 1, 3, 6, 12, and 24mo after I-CXL. RESULTS: After 24mo I-CXL, compared to the level at baseline, BCVA significantly improved 0.14±0.07 (P=0.010); mean keratometry signifi­cantly decreased 0.72±1.97 (P=0.021); maximum keratometry significantly reduced 2.30±5.01 (P=0.014); central keratoconus index significantly reduced 0.04±0.08 (P=0.007). The demarcation line was visible in 83.1% of eyes at 1mo after treatment, with a depth of 298.95±51.97 μm, and gradually indistinguishable. One eye had repeat treatment. Intraocular pressure and endothelial cell density did not change significantly. CONCLUSION: I-CXL using 0.1% riboflavin halts keratoconus progression within 24mo, resulting in a significant improvement in visual and topographic parameters. Moreover, the depth of the demarcation line is similar to that previously reported in standard epithelium-off CXL procedures.

Background: Ginseng is widely used as herb or food. Different parts of ginseng have diverse usages. However, the comprehensive analysis on the ginsenosides in different parts of ginseng root is scarce. Methods: An ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) combined with UNIFI informatics platform and ultra-high-performance liquid chromatography-charged aerosol detection (UHPLC-CAD) were employed to evaluate the different parts of cultivated ginseng root. Results: 105 ginsenosides including 16 new compounds were identified or tentatively characterized. 22 potential chemical markers were identified, 20, 17, and 19 for main root (MR) and fibrous root (FR), main root (MR) and branch root (BR), and main root (MR) and rhizome (RH), respectively. The relative contents of Re, Rb1, 20(R)-Rh1, Rd, and Rf were highest in FR. The relative content of Rg1 was highest in RH. The total relative content of pharmacopoeia indicators Rg1, Re, and Rb1 was highest in FR. Conclusion: The differences among these parts were the compositions and relative contents of ginsenosides. Under our research conditions, the peak area ratio of Rg1 and Re could distinguish the MR and FR samples. Fibrous roots showed rich ingredients and high ginsenosides contents which should be further utilized.

Signal transducer and activator of transcription 3 (STAT3) is an attractive therapeutic target for cancer treatment. In this study, we identify lycorine is an effective inhibitor of STAT3, leading to repression of multiple oncogenic processes in colon carcinoma. Lycorine selectively inactivates phospho-STAT3 (Tyr-705), and subsequent molecular docking uncovers that lycorine directly binds to the SH2 domain of STAT3. Consequently, we find that lycorine exhibits anti-proliferative activity and induces cell apoptosis on human colorectal cancer (CRC) in vitro. Lycorine induces the activation of the caspase-dependent mitochondrial apoptotic pathway, as indicated by activation of caspase and increase of the ratio of Bax/Bcl-2 and mitochondrial depolarization. Overexpressing STAT3 greatly blocks these effects by lycorine in CRC cells. Finally, lycorine exhibits a potential therapeutic effect in xenograft colorectal tumors by targeting STAT3 without observed toxicity. Taken together, the present study indicates that lycorine acts as a promising inhibitor of STAT3, which blocks tumorigenesis in colon carcinoma.

AIM:To evaluate the enrichment of riboflavin in the corneal stroma after intracameral injection to research the barrier ability of the corneal endothelium to riboflavin in vivo.METHODS:The right eyes of 30 New Zealand white rabbits were divided into three groups. Different concentrations riboflavin-balanced salt solutions (BSS) were injected into the anterior chamber (10 with 0.5%, 10 with 1%, and 10 with 2%). Eight corneal buttons of 8.5 mm in diameter from each group were dissected at 30min after injection and the riboflavin concentrations in the corneal stroma were determined using high-performance liquid chromatography (HPLC) after removing the epithelium and endothelium. The other two rabbits in every group were observed for 24h and sacrificed. As a comparison, the riboflavin concentrations from 16 corneal stromal samples were determined using HPLC after instillation of 0.1% riboflavin-BSS solution for 30min on the corneal surface (8 without epithelium and 8 with intact epithelium).RESULTS: The mean riboflavin concentrations were 11.19, 18.97, 25.08, 20.18, and 1.13 µg/g for 0.5%, 1%, 2%, de-epithelialzed samples, and the transepithelial groups, respectively. The color change of the corneal stroma and the HPLC results showed that enrichment with riboflavin similar to classical de-epithelialized corneal collagen crosslinking (CXL) could be achieved by intracameral1% riboflavin-BSS solution after 30min; the effect appeared to be continuous for at least 30min.CONCLUSION:Riboflavin can effectively penetrate the corneal stroma through the endothelium after an intracameral injection in vivo, so it could be an enhancing method that could improve the corneal riboflavin concentration in transepithelial CXL.

AIM: To report a two-year's results of iontophoresis-assisted transepithelial corneal cross-linking(I-CXL)for progressive keratoconus. METHODS: Thirty-four eyes in 24 patients with progressive keratoconus(mean age 21.0±5.6 years; range: 14-32 years)were treated. After 1g/L riboflavin-distilled water solution was administered by iontophoresis-assited(current 1mA)transepithelial method for 5min in total, standard surface UVA irradiation(370nm, 3mW/cm2)was performed at a 1-cm distance for 30min. The best corrected visual acuity(BCVA)measured as LogMAR number, corneal refractive astigmatism, K1, K2, Kmean, Kmax, intraocular pressure, endothelial cell density, the thickness at corneal apex and the thinnest point were measured preoperatively and 2a postoperatively. RESULTS:At 2a after the procedure, BCVA(LogMAR)improved from 0.32±0.25 to 0.25±0.19(t=2.849, P=0.015). K1 decreased from 47.12±4.33 to 46.06±4.77(t=2.652, P=0.015). K2 decreased from 51.36±5.59 to 50.40±6.16(t=2.121, P=0.047). Kmean decreased from 49.12±4.76 to 48.10±5.25(t=2.663, P=0.015). Kmax decreased from 57.57±8.30 to 55.91±8.14(t=2.398, P=0.026). The corneal apex thickness decreased from 476.90±38.71μm to 454.43±40.86μm(t=2.853, P=0.010). The thinnest thickness decreased from 464.38±39.92μm to 433.86±50.78μm(t=3.485, P=0.002). Corneal refractive astigmatism, intraocular pressure and endothelial cell density did not show significant changes. CONCLUSION: I-CXL for progressive keratoconus is safe and effective which can prevent deterioration of progressive keratoconus within 2a, but further long-term studies are necessary still.

Eleven new furostanol saponins, typaspidosides B-L (1-11), one new spirostanol saponin, typaspidoside M (12), and five known spirostanol saponins, 25S-atropuroside (13), neoaspidistrin (14), (25S)-pratioside D1 (15), 25S-aspidistrin (16) and 25S-neosibiricoside (17) were isolated from the rhizomes of Aspidistra typica Baill. The structures of the new compounds were established using 1D and 2D NMR (1H-1H COSY, HMQC, HMBC and ROESY) spectroscopy, high resolution mass spectrometry, and chemical methods. The aglycones of 1-3 (unusual furostanol saponins with opened E ring type), 9 and 10 (the methoxyl substituent at C-23 position) were found, identified from natural products for the first time. Moreover, the anti-HIV activities of the isolated steroidal glycosides were assessed, and compounds 13, 14, 16 and 17 exhibited high active against HIV-1.

AIM: To compare the difference in measurements of central corneal thickness(CCT)using A-scan,corneal specular microscopy and Pentacam in keratoconus. METHODS: Between July 2012 and October 2012, the CCT of 31 patients(55 eyes)were measured by A-scan, corneal specular microscopy and Pentacam. The results were analyzed by F-test and Pearson correlation. RESULTS: The values of CCT measured by A-scan(55 eyes), corneal specular microscopy(45 eyes)and Pentacam(52 eyes)were 469.87±57.56, 479.00±42.39, and 487.02±44.64μm, respectively; F-test results showed there were no statistical significant differences between CCT measured by three different instruments(P>0.05). The correlation between the measurements was evaluated using Pearson correlation coefficients. The CCT values by these three instruments were positively related by linear correlation analysis, A-scan and Pentacam measurements(r1=0.758, Pr2=0.949, Pr3=0.685, PCONCLUSION: There is a high correlation between these three instruments. A-scan and Pentacam are more precise than corneal specular microscopy. Pentacam system can measure the CCT easily, accurately and without any invasion. It is more suitable for people with keratoconus to monitor the every point of cornea and for people to do more deep research.

The thermodynamic equilibrium of deoxidation reactions between molten slag and steel was calculated using a slag-steel coupling thermodynamic model and the mass conservation model based on the ion-molecular coexistence theory. The study focused on the effects of slag composition and deoxidizer type on the oxygen content of low alloy steel during the electroslag remelting (ESR) process. The measured and predicted values of the oxygen content in remelted ingots, and the contents of FeO and MnO in slags were compared and analyzed. Results show that the measured content of total oxygen has a certain correlation with the trend of dissolved oxygen predicted by the model when using Ca-Si alloys as deoxidizer, but it is not correlated with the trend of dissolved oxygen predicted by the model when using Al as deoxidizer. The deoxidation mechanisms of Ca-Si and Al are different. Ca-Si alloy directly reacts with FeO and MnO in slag to reduce the oxygen potential of slag, hence it can inhibit the transfer of oxygen from the slag to molten steel. While, when Al deoxidizer is used, the oxygen content in steel is mainly reduced through floating up the alumina inclusions. Compared to Al, utilizing Ca-Si alloy as a deoxidizer is more effective in reducing the oxygen content and the amount of inclusions in ESR ingot. [ABSTRACT FROM AUTHOR]