Improved glucose yield and concentration of sugarcane bagasse by the pretreatment with ternary deep eutectic solvents and recovery of the pretreated liquid.

In this study, a novel ternary deep eutectic solvents (DES) consisting of choline chloride/PEG/hydroxyethyl sulfonic acid (HSA) was developed to effectively improve glucose yield and concentration of sugarcane bagasse, and the conditions of the pretreatment were optimized by response surface method (RSM). Under the optimal conditions, the maximum glucose concentration (GC) could reach 12.39 g/L (HSA concentration 1.34 %, PEG400, 2.3 h, 150 °C), and the maximum glucose yield (GY) was 0.2497 g/g (HSA concentration 1.41 %, PEG400, 2.1 h, 150 °C). Hemicellulose was completely removed, and the maximum lignin removal rate was 86.89 %. After pretreatment, 95 % of the pretreated liquid can be recycled. Finally, the structural and morphological changes of bagasse before and after pretreatment were investigated by scanning electron microscopy (SEM), Fourier Transform infrared analyzer (FT-IR) and X-ray diffraction (XRD).

An Automatic Immunoaffinity Pretreatment of Deoxynivalenol Coupled with UPLC-UV Analysis.

An immunoaffinity magnetic beads (IMBs) based automatic pretreatment method was developed for the quantitative analysis of deoxynivalenol (DON) by ultra-performance liquid chromatography and ultraviolet detector (UPLC-UV). First, N-hydroxysuccinimide-terminated magnetic beads (NHS-MBs) with good magnetic responsivity and dispersibility were synthesized and characterized by optical microscopy, scanning electron microscopy (SEM), and laser diffraction-based particle size analyzer. Then, the amino groups of anti-DON monoclonal antibody (mAb) and the NHS groups of NHS-MBs were linked by covalent bonds to prepare IMB, without any activation reagent. The essential factors affecting the binding and elution of DON were meticulously tuned. Under optimal conditions, DON could be extracted from a real sample and eluted from IMB by water, enabling environmentally friendly and green analysis. Hence, there was no need for dilution or evaporation prior to UPLC-UV analysis. DON in 20 samples could be purified and concentrated within 30 min by the mycotoxin automated purification instrument (MAPI), allowing for automated, green, high-throughput and simple clean-up. Recoveries at four distinct spiking levels in corn and wheat ranged from 92.0% to 109.5% with good relative standard deviations (RSD, 2.1-7.0%). Comparing the test results of IAC and IMB in commercial samples demonstrated the reliability and superiority of IMB for quantitatively analyzing massive samples.

Vacuum-assisted black liquor-recycling enhances the sugar yield of sugarcane bagasse and decreases water and alkali consumption.

Black liquor (BL) remains a critical problem during alkaline pretreatment. To solve this issue, a novel pretreatment strategy termed vacuum-assisted black liquor-recycling pretreatment, was established to pretreat sugarcane bagasse (SCB). Firstly, SCB was pretreated with 2% NaOH at 121 °C for 1 h under vacuum conditions. The produced BL was used for subsequent pretreatments after pH recovery with NaOH. The pretreated SCBs were subject to enzymatic hydrolysis and separate hydrolyzation and fermentation (SHF) without washing to neutral pH. BL was recycled on seven occasions. The results indicated that glucose yields did not significantly differ between pretreatment with NaOH and recovered BL. The enzymatic hydrolysis and the fermentation resulted in maximum 0.35 g/g of glucose yield and 116.5 g/kg of ethanol yield respectively. Compared with conventional pretreatment with NaOH, the VABLR method showed high conversion rates of cellulose into monosaccharaides, whilst preserving ~20% and ~46% of alkali and water usage, respectively.

Pre-treatment of sugarcane bagasse with aqueous ammonia-glycerol mixtures to enhance enzymatic saccharification and recovery of ammonia.

In this work, an efficient aqueous ammonia with glycerol (AAWG) method to improve the digestibility of sugarcane bagasse (SCB) was developed. Response surface methodology was utilized to optimize the AAWG parameters to achieve the maximum total fermentable sugar concentration (TFSC) and total fermentable sugar yield (TFSY). Under optimal AAWG conditions, 13.59 g/L TFSC (9.25% ammonia, 1.86 h, 180 °C) and 0.4449 g/g TFSY (9.51% ammonia, 1.78 h, 180 °C) were achieved, with delignification of 77.81% and 70.91%, respectively. Compared to pretreatment with glycerol or aqueous ammonia, the AAWG method significantly enhanced the enzymatic efficiency of SCB. The ammonia was recovered from the pretreatment liquid by distillation, and about one-third of the ammonia was retained. The overall results indicate that AAWG is effectively used as a pretreatment method for recovering ammonia, which would largely contribute to the economic benefits of biomass biorefinery.

Enhanced enzymatic saccharification of sugarcane bagasse pretreated by sodium methoxide with glycerol.

Sodium methoxide (CH3ONa) with glycerol pretreatment (CWGP) was performed to improve the enzymatic digestibility of sugarcane bagasse (SCB). Response surface methodology was utilized to optimize the CWGP parameters for pretreating SCB from the perspective of total fermentable sugar yield (TFSY) and total fermentable sugar concentration (TFSC). Under the optimal CWGP conditions, 0.5666g/g of TFSY (0.82% CH3ONa, 1.11h, 150°C) and 17.75g/L of TFSC (0.87% CH3ONa, 1.38h, 149.27°C) were achieved, corresponding to delignification of 79.05% and 79.34%, respectively. Compared the pretreatment using glycerol or CH3ONa alone, the CWGP has significant synergies to enhance the enzymatic efficiency of SCB. The physical and chemical characteristics of untreated and pretreated SCBs were analyzed using FT-IR, XRD, and SEM, and the results suggest that CWGP significantly increased the susceptibility of the substrates to enzymatic digestibility. Ultimately, CWGP might be a prospective candidate for the pretreatment process of enzyme-based lignocellulosic biorefineries.

Vacuum-assisted alkaline pretreatment as an innovative approach for enhancing fermentable sugar yield and decreasing inhibitor production of sugarcane bagasse.

Sodium hydroxide pretreatment of sugarcane bagasse under vacuum conditions was established and evaluated in this study. Compared to pretreatment under conventional moderate pressure conditions, only half of the total phenolic compounds and less than half of the formic acid were produced under vacuum conditions, while the yield of total fermentable sugar was significantly increased by 31.38%. The pretreatment parameters: NaOH concentration, pretreatment time, and pretreatment temperature, were optimized using response surface methodology based on the response values of the total fermentable sugar yield (TFSY) and the total fermentable sugar concentration (TFSC), respectively. Under the optimal conditions, the TFSY of 0.5146g/g and the TFSC of 17.37g/L were achieved, respectively. By adjusting the ratio of cellulases to xylanase, the TFSY reached a maximum of 0.5213g/g when the ratio was 1:1, while the maximum TFSC of 17.71g/L was achieved when the ratio was 1:4.

Comparative study of different alcoholate pretreatments for enhanced enzymatic hydrolysis of sugarcane bagasse.

Pretreatment of sugarcane bagasse (SCB) with alcoholates, sodium methoxide (CH3ONa), potassium methoxide (CH3OK) and sodium ethoxide (C2H5ONa), was investigated. Analyses of lignocellulose composition and enzymatic saccharification indicated that C2H5ONa showed the highest enzymatic efficiency of 102.1%. The response surface optimization of C2H5ONa pretreatment showed that under optimal conditions (4% of C2H5ONa, 121°C, 1h), 65.4% of lignin was removed and the enzymatic efficiency reached 105.2%. Hydrolysis of SCB with cellulases and xylanase at a ratio of 4:1 showed the strongest synergism with reducing sugar production of 21g/L and conversion rates of cellulose and xylan reaching 110.4% and 94.5%, respectively. These results indicated that C2H5ONa is a promising alkali to pretreat SCB and the synergism between cellulases and xylanase has a significant effect on enzymatic saccharification of the pretreated SCB.

Cucurbitacin E Induces Autophagy via Downregulating mTORC1 Signaling and Upregulating AMPK Activity.

Cucurbitacins, the natural triterpenoids possessing many biological activities, have been reported to suppress the mTORC1/p70S6K pathway and to induce autophagy. However, the correlation between such activities is largely unknown. In this study, we addressed this issue in human cancer cells in response to cucurbitacin E (CuE) treatment. Our results showed that CuE induced autophagy as evidenced by the formation of LC3-II and colocalization of punctate LC3 with the lysosomal marker LAMP2 in HeLa and MCF7 cells. However, CuE induced much lower levels of autophagy in ATG5-knocked down cells and failed to induce autophagy in DU145 cells lacking functional ATG5 expression, suggesting the dependence of CuE-induced autophagy on ATG5. Consistent with autophagy induction, mTORC1 activity (as reflected by p70S6K and ULK1S758 phosphorylation) was inhibited by CuE treatment. The suppression of mTORC1 activity was further confirmed by reduced recruitment of mTOR to the lysosome, which is the activation site of mTORC1. In contrast, CuE rapidly activated AMPK leading to increased phosphorylation of its substrates. AMPK activation contributed to CuE-induced suppression of mTORC1/p70S6K signaling and autophagy induction, since AMPK knockdown diminished these effects. Collectively, our data suggested that CuE induced autophagy in human cancer cells at least partly via downregulation of mTORC1 signaling and upregulation of AMPK activity.

Association of Toll-like receptor 7 and 8 gene polymorphisms with Graves' disease in Chinese Cantonese population.

Graves' disease (GD) is a common polygenic multifactorial autoimmune disease. Toll-like receptors (TLRs) play critical roles in the activation of innate and adaptive immune responses. This study investigated the association of TLR7 and TLR8 gene polymorphisms with susceptibility of GD. Five single nucleotide polymorphisms (SNPs), namely, rs179019, rs179010 and rs3853839 in TLR7 and rs3764880 and rs5744088 in TLR8, were evaluated in 332 GD patients and 351 controls using High-Resolution Melting analysis. After adjusting for age, SNP rs179010 was found to decrease the risk of GD in females (OR(T vs C) = 0.64, P = 0.004). In the additive model, the risk of GD decreased significantly as the number of T alleles increased in females [odds ratio (OR) = 0.67 (0.50-0.90), P = 0.007]. The multivariate logistic regression analysis confirmed the independent contribution of rs179010 to the protective effect against GD. This study indicates that rs179010 in TLR7 may be associated with the decreased susceptibility to GD in Chinese Cantonese.

Polymorphisms in TLR1, TLR6 and TLR10 genes and the risk of Graves' disease.

Graves' disease (GD) is postulated to be caused by the combined effects of susceptibility genes and environmental triggers. Toll-like receptors (TLRs) play a role in the activation of innate and adaptive immune responses in mammalians. The aim of this study was to evaluate the potential association of polymorphisms in TLR1, TLR6 and TLR10 genes with GD in Chinese Cantonese population. Seven single nucleotide polymorphisms (i.e. rs4833095 and rs5743565 in TLR1; rs5743808 in TLR6; and rs4504265, rs11466655, rs11096957 and rs10856839 in TLR10) were evaluated in 332 GD patients and 351 unrelated controls from Chinese Cantonese population. SNP rs5743565 in TLR1 conferred a protective effect against GD. The minor allele G of rs5743565 decreased the risk of GD in all cases (odds ratio; ORG vs. A=0.72 (0.58-0.91); p=0.005; ptrend=0.004) and early onset patients (ORG vs. A=0.72 (0.56-0.91); p=0.007; ptrend=0.006). This study provided evidence that genetic variation rs5743565 in TLR1 might be associated with the decreased susceptibility of GD.

Direct fermentation of amorphous cellulose to ethanol by engineered Saccharomyces cerevisiae coexpressing Trichoderma viride EG3 and BGL1.

Direct ethanol fermentation from amorphous cellulose was achieved using an engineered industrial Saccharomyces cerevisiae strain. Two cellulase genes endoglucanase (eg3) and ß-glucosidase (bgl1) were obtained from Trichoderma viride and integrated into the genome of S. cerevisiae. These two cellulases could be constitutively coexpressed and secreted by the recombinant strain S. cerevisiae-eb. The enzyme activities were analyzed in the culture supernatants, with the highest endoglucanase activity of 2.34 units/ml and ß-glucosidase activity of 0.95 units/ml. The effects of pH, temperature and metal ions on enzyme activities were analyzed. The coexpression strain S. cerevisiae-eb could grow in carboxymethyl cellulose (CMC) and utilize it as the single carbon source. The 20 g/L CMC as a model substrate of amorphous cellulose was used in fermentation. The ethanol production reached 4.63 g/L in 24 h, with the conversion ratio of 64.2% compared with the theoretical concentration. This study demonstrated that the engineered industrial strain S. cerevisiae-eb could convert amorphous cellulose to ethanol simultaneously and achieve consolidated bioprocessing (CBP) directly.

Association of TLR4 and TLR5 gene polymorphisms with Graves' disease in Chinese Cantonese population.

Graves' disease (GD) is postulated to be caused by the combined effects of susceptibility genes and environmental triggers. Toll like receptors (TLRs) play a role in the activation of innate and adaptive immune responses in mammalians. The aim of this study was to evaluate the potential association of TLR4 and TLR5 gene polymorphisms with GD in Chinese Cantonese population. Four single nucleotide polymorphisms (SNPs), rs11536889 and rs7873784 in TLR4, rs2072493 and rs5744174 in TLR5, were evaluated in 332 GD patients and 351 unrelated controls from Chinese Cantonese population. The minor allele C of TLR5 rs5744174 decreased the risk to GD in females (ORC vs. T=0.63; p=0.003; ptrend=0.003). Under a dominant model, rs5744174 conferred a protective effect in all cases (ORCC/CT vs. TT=0.65; p=0.009) or female subset (ORCC/CT vs. TT=0.57; p=0.002). Under a co-dominant model, rs5744174 also conferred a protective effect in all cases (ORTC vs. TT=0.64; p=0.008) and females (ORTC vs. TT=0.57; p=0.002). The haplotype A-C of TLR5 (rs2072493-rs5744174) decreased the risk of GD in females (OR=0.62; p=0.002). The other three SNPs were not found associated with GD. This study provided evidence that polymorphisms in TLR5 might be associated with decreased susceptibility of GD in females.

Ethanol production from xylan-removed sugarcane bagasse using low loading of commercial cellulase.

Xylan was always extracted as the feedstock for xylooligosaccharides production. The xylan-removed residue may contain high content of cellulose and thus had a possibility to be converted into ethanol. After soaked in 12% of NaOH at room temperature overnight, solubilization of cellulose, xylan, and lignin was 4.64%, 72.06%, and 81.87% respectively. The xylan-removed sugarcane bagasse (XRSB) was enzymatically hydrolyzed by using decreased cellulase loadings. The results showed that 7.5 FPU/g cellulose could obtain a cellulose conversion yield of 82%. Increasing the cellulase loading did not result in higher yield. Based on this, bioethanol production was performed using 7.5 FPU/g cellulose by employing fed-batch fermentation mode. The final ethanol concentration reached 40.59 g/L corresponding to 74.2% of the theoretical maximum. The high titer ethanol and low cellulase loading may reduce the overall cost.

One-pot simultaneous saccharification and fermentation: a preliminary study of a novel configuration for cellulosic ethanol production.

Combination of size reduction and mild alkali pretreatment may be a feasible way to produce bioethanol without rinsing and detoxifying the solid substrate. Based on that, a fermentation configuration named one-pot SSF in which pretreatment and fermentation steps were integrated was developed. Additionally, the effect of laccase on fermentation performance was investigated. Delignification was the major effect of the alkali pretreatment at 121°C for 60min. The highest glucose and xylose yield, which obtained from the smallest particle at a substrate loading of 2%, was 6.75 and 2.71g/L, respectively. Laccase improved the fermentation efficiency by 6.8% for one-pot SSF and 5.7% for SSF. Bioethanol from one-pot SSF with laccase supplementation reached 67.56% of the theoretical maximum, whereas that from SSF with laccase supplementation reached 57.27%. One-pot SSF might be a promising configuration to produce bioethanol because of 100% solid recovery, and rinsing water and detoxification elimination.

Synergism of cellulase, xylanase, and pectinase on hydrolyzing sugarcane bagasse resulting from different pretreatment technologies.

Sugarcane bagasse (SCB) resulting from different pretreatments was hydrolyzed by enzyme cocktails based on replacement of cellulase (Celluclast 1.5 L:Novozym 188=1FPU:4pNPGU) by xylanase or pectinase at different proportions. Lignin content of NaOH pretreated SCB and hemicellulose content of H2SO4 pretreated SCB were the lowest. NaOH pretreatment showed the best for monosaccharide production among the four pretreatments. Synergism was apparently observed between cellulase and xylanase for monosaccharide production from steam exploded SCB (SESB), NaOH, and H2O2 pretreated SCB. No synergism was observed between cellulase and pectinase for producing glucose. Additionally, no synergism was present when H2SO4 pretreated SCB was used. Replacement of 20% of the cellulase by xylanase enhanced the glucose yield by 6.6%, 8.8%, and 9.5% from SESB, NaOH, and H2O2 pretreated SCB, respectively. Degree of synergism between cellulase and xylanase had positive relationship with xylan content and was affected by hydrolysis time.

[Association of P53 gene polymorphisms with susceptibility to endometriosis].

OBJECTIVE: To assess the association of a single nucleotide polymorphism(SNP) in tumor suppressor gene P53 with the risk of endometriosis (EM) in Han Chinese women. METHODS: For 460 EM patients, 113 patients with endometrial carcinoma and 530 matched unrelated controls, a rs1042522(C/G) SNP of the P53 gene was genotyped by polymerase chain reaction-single strand polymorphism (PCR-SSP) and DNA sequencing. RESULTS: A significant difference has been detected in the distribution of rs1042522 alleles and genotypes between the EM patients and controls (P< 0.01). Allele G has increased the risk of EM by 1.209 times, while allele C has reduced this risk by 0.837 times. Compared with GG genotype, GC and CC genotypes have both increased the risk for EM (OR=2.073, 95%CI: 1.521-2.820, and OR=1.930, 95%CI: 1.363-2.733, respectively). Significant differences were also detected in the distribution of rs1042522 alleles and genotypes between endometrial carcinoma patients and controls (P< 0.01). Allele G has increased the risk to endometrial carcinoma by 1.311 times, while allele C has reduced this risk by 0.757 times. Compared with GG genotypes, individuals with GC and CC genotypes are more likely to be affected with endometrial carcinoma (OR=2.778, 95%CI: 1.585-4.870, and OR=2.864, 95%CI: 1.557-5.263, respectively). CONCLUSION: Our study has suggested a significant association between the rs1042522(G/C) polymorphism and susceptibility to EM in Han Chinese women. The mechanism of EM is similar to carcinoma from genetics point of view.

Solvent extraction of antioxidants from steam exploded sugarcane bagasse and enzymatic convertibility of the solid fraction.

Solvent extraction of steam exploded lignocellulosic biomass may be a potential way to obtain antioxidative extracts and to enhance the enzymatic convertibility of the solid residue. Boiling solvent extraction (BSE) showed higher solid and phenolic yields than room temperature extraction. Solubilities of phenolics and sugars were higher in anhydrous ethanol (AE) and deionized water (DW) than in ethyl acetate under each individual extraction condition. The antioxidant activities of the AE and DW extract obtained under BSE were better than those of 10mM vitamin C. Conversion of the solid fractions into reducing sugar using Celluclast 1.5L and Novozym 188 after AE and DW extraction was 95.13% and 92.97%, respectively, higher than that obtained with SESB (88.95%).

Identification and characterization of a novel thermostable gh-57 gene from metagenomic fosmid library of the Juan de Fuca Ridge hydrothemal vent.

A novel glycoside hydrolases family 57 gene (gh-57) was found from a metagenomic fosmid library constructed from a black smoker chimney sample 4143-1 from the Mothra hydrothermal vent at the Juan de Fuca Ridge. Sequence and homology analysis using BLAST revealed that it had high similarity to gh-57 family. Conserved domain research revealed that the novel gh-57 contained a Glyco-hydro-57 domain and five conserved regions, including two putative catalytic residues Glu¹54 and Asp²6³. The three-dimensional features of the protein and its homologue from Pyrococcus horikoshii OT3 known as α-amylase were generated by homology modeling. The gh-57 gene was cloned, expressed, and purified in Escherichia coli using pQE system. Enzyme activity revealed that the recombinant protein could hydrolyze soluble starch and demonstrated amylase activity. It showed an optimal pH of 7.5, an optimal temperature of 90 °C, and its thermostability at 90 °C could remain over 50% enzyme activity for 4 h. The enzyme activity could be increased by DTT and Mg²âº while an inhibitory effect was observed with EDTA, ATP, and Ca²âº. These results showed that the gh-57 gene was a novel thermostable amylase from oceanic microorganisms.

HLA class II polymorphisms associated with the physiologic characteristics defined by Traditional Chinese Medicine: linking modern genetics with an ancient medicine.

OBJECTIVES: The aim of this study was to test whether human leukocyte antigen (HLA) polymorphism contributes to the physical constitutions classified in Traditional Chinese Medicine (TCM). DESIGN: Seven hundred six (706) individuals of the Han ethnic group inhabiting South China were classified into 7 TCM constitution groups, according to the criteria described in Theories of Physical Constitutions of Traditional Chinese Medicine, and the distributions of HLA-DRB1, DPB1, and DQB1 were investigated using the polymerase chain reaction-sequencing-based typing method. RESULTS: The allele frequencies of DPB1*0501 in the Yin-deficiency group, DRB1*09012 in the Phlegm-wetness group, and DQB1*03032 in the Qi-deficiency and Phlegm-wetness groups were significantly different from that of the corresponding alleles in the Normality constitution, suggesting those alleles might be group-specific alleles and thus related to a particular constitution. Based on our analysis of serological groups of HLA, the associations of DR*04 with the Blood-stasis group and DQ*09 with the Qi-deficiency and Phlegm-wetness groups were observed. CONCLUSIONS: This was the first study to systematically investigate the relationship between HLA and TCM constitution using a high-resolution typing technique. The results suggested a genetic basis for the classification of physical constitution in TCM. This study laid the foundation, for the first time ever, toward gaining insight into the theory of traditional medicine using modern biological approaches.

Acute phase response in zebrafish upon Aeromonas salmonicida and Staphylococcus aureus infection: striking similarities and obvious differences with mammals.

Zebrafish has emerged as a valuable model for immunological studies. However, little is known about the overall picture of its immune response to infectious pathogens. Here we present the first systematic study of its immune response to Aeromonas salmonicida and Staphylococcus aureus, a Gram-negative and a Gram-positive bacteria, respectively. Genes induced upon infection were identified with suppression subtractive hybridization, with many of them encoding acute phase proteins (APPs). When compared with mammals, striking similarities and obvious differences have been observed. Both similar APPs (SAA, hepcidin and haptoglobin, etc.) and a similar system for the induction of APPs (which involves the TLRs, pro-inflammatory cytokines and C/EBPs) were identified, implying evolutionary conserved mechanisms among fish and mammals. Some novel APPs were also discovered, suggesting different immune strategies adopted by fish species. Among which, LECT2 was induced by up to 1000-fold upon infection, shedding new lights on the function of this gene. Our results constitute the first demonstration of a similar while different immune response in zebrafish and open new avenues for the investigation of evolutionary conserved and fish specific mechanisms of innate immunity.