Production and purification of 2-phenylethanol by Saccharomyces cerevisiae using tobacco waste extract as a substrate

2-phenylethanol (2-PE), which is extracted naturally from plant or biotechnology processing, is widely used in the food and cosmetics industries. Due to the high cost of 2-PE production, the valorization of waste carbon to produce 2-PE has gained increasing attention. Here, 2-PE was produced by Saccharomyces cerevisiae using tobacco waste extract (TWE) as the substrate. Considering the toxicity of nicotine and its inhibition of 2-PE, the tolerance of S. cerevisiae was first evaluated. The results suggested that the production of 2-PE by S. cerevisiae in TWEs could be carried out at 2·0 mg ml⁻¹ nicotine concentrations and may be inhibited by 1·0 mg ml⁻¹ 2-PE. Thus, the compounds in the TWEs prepared at different temperatures were detected, and the results revealed that the TWEs prepared at 140°C contained 2·18 mg ml⁻¹ of nicotine, had total sugar concentrations of 26·8 mg ml⁻¹ and were suitable for 2-PE production. Due to feedback regulation, the 2-PE production was only 1·11 mg ml⁻¹, and the remaining glucose concentration remained at 13·78 mg ml⁻¹, which indicated insufficient glucose utilization. Then, in situ product recovery was further implemented to remove this inhibition; the glucose utilization (the remaining concentration decreased to 3·64 mg ml⁻¹) increased, and the 2-PE production increased to 1·65 mg ml⁻¹. The 2-PE produced in the fermentation broth was first isolated by elution from the resin with 75% ethanol and then by removing the impurities with 2·5% activated charcoal, and pure 2-PE was identified by gas chromatography mass spectrometry. The results of this study suggest that TWE could be an alternative carbon source for 2-PE production. This could provide an outlet tobacco waste as well as reducing the price of natural 2-PE, although more strategies need to be explored to improve the production yield of 2-PE by using TWE.     

2-phenylethanol (rose aroma) production potential of an isolated pichia kudriavzevii through solid-state fermentation

2-phenylethanol (2-PE) is a higher alcohol widely used in industry that can be obtained by solid-state fermentation (SSF) using low-cost raw materials. This report describes the 2-PE production potential of an indigenous Pichia kudriavzevii isolated from solid-state fermented sugarcane bagasse that possesses attractive characteristics for processing waste streams such as its low-pH tolerance, high growth rate and temperature resistance. Besides, 2-PE production was optimized in batch-SSF using sugarcane bagasse supplemented with l-phenylalanine as substrate. Full factorial design allowed identifying the pH adjustment, micronutrient addition, inoculum and co-substrate load effects, and response surface methodology served to identify the maximum production based on temperature, initial moisture content (MC₀) and specific airflow rate (S_AFR). While the pH adjustment and micronutrient addition did not affect the 2-PE production, temperature and MC₀ resulted critical for the process. After optimization, the maximum 2-PE content was 27.2 ± 0.2 mg per gram of dry substrate at 31 °C, 76 % MC₀ and 0.129 L h⁻¹ g⁻¹ S_AFR. This result was 23.8 % higher than the sub-optimal condition, and it is the highest 2-PE production via SSF reported so far. These results confirm the ability of P. kudriavzevii for producing 2-PE, and its potential for using waste streams as substrate.     

Biosorption of sterols from tobacco waste extract using living and dead of newly isolated fungus Aspergillus fumigatus strain LSD-1

ABSTRACT

Sterols are verified to be able to produce polycyclic aromatic hydrocarbons during its pyrolysis. In this study, a kind of Aspergillus fumigatus (LSD-1) was isolated from cigar leaves, and the biosorption effects on the stigmasterol, β-sitosterol, campesterol, cholesterol, and ergosterol by using living and dead biomass of LSD-1 were investigated. The results showed that both living and dead biomass could efficiently remove these sterols in aqueous solution and tobacco waste extract (TWE). Interestingly, compared with the living biomass of LSD-1, the dead biomass of LSD-1 not only kept a high adsorption efficiency but also did not produce ergosterol. Overall, dead biomass of LSD-1 was a more suitable biosorbent to sterols in TWE. Furthermore, Brunner-Emmet-Teller (BET), Fourier transformed infrared spectrometer (FTIR) and scanning electron microscope (SEM) analysis were used to explore the biosorption process of living and dead biomass and their differences, suggesting that the biosorption of sterols was a physical process.     

Biodegradation of nicotine by a newly isolated Agrobacterium sp. strain S33

Aims: To isolate and characterize bacteria capable of degrading nicotine from the rhizospheric soil of a tobacco plant and to use them to degrade the nicotine in tobacco solid waste. Methods and Results: A bacterium, strain S33, was newly isolated from the rhizospheric soil of a tobacco plant, and identified as Agrobacterium sp. based on morphology, physiological tests, Biolog MicroLog3 4·20 system and 16S rRNA gene sequence. Using nicotine as the sole source of carbon and nitrogen in the medium, it grew optimally with 1·0 g l^?1 of nicotine at 30°C and pH 7·0, and nicotine was completely degraded within 6 h. The resting cells prepared from the glucose‐ammonium medium or LB medium could not degrade nicotine within 10 h, while those prepared from the nicotine medium could completely degrade 3 g l^?1 of nicotine in 1·5 h at a maximal rate of 1·23 g nicotine h^?1 g^?1 dry cell. Using the medium containing nicotine, glucose and ammonium simultaneously to cultivate strain S33, the resting cells could degrade 98·87% of nicotine in tobacco solid waste with the concentration as 30 mg nicotine g^?1 dry weight tobacco solid waste within 7 h at a maximal rate of 0·46 g nicotine h^?1 g^?1 dry cell. Conclusions: This is the first report that Agrobacterium sp. has the ability to degrade nicotine. Agrobacterium sp. S33 could use nicotine as the sole source of carbon and nitrogen. The use of resting cells of the strain S33 prepared from the nicotine–glucose–ammonium medium was an effective method to degrade nicotine and detoxify tobacco solid waste. Significance and Impact of the Study: Nicotine in tobacco wastes is both toxic and harmful to human health and the environment. This study showed that Agrobacterium sp. S33 may be suitable for the disposal of tobacco wastes and reducing the nicotine content in tobacco leaves.     

3D pharmacophore models for thromboxane A2 receptor antagonists

Thromboxane A₂ (TXA₂) is an endogenous arachidonic acid derivative closely correlated to thrombosis and other cardiovascular diseases. The action of TXA₂ can be effectively inhibited with TXA₂ receptor antagonists (TXRAs). Previous studies have attempted to describe the interactions between the TXA₂ receptor and its ligands, but their conclusions are still controversial. In this study, ligand-based computational drug design is used as a new and effective way to investigate the structure-activity relationship of TXRAs. Three-dimensional pharmacophore models of TXRAs were built with HypoGenRefine and HipHop modules in CATALYST software. The optimal HypoGenRefine model was developed on the basis of 25 TXRAs. It consists of two hydrophobic groups, one aromatic ring, one hydrogen-bond acceptor and four excluded volumes. The optimal HipHop model contains two hydrophobic groups and two hydrogen-bond acceptors. These models describe the key structure-activity relationship of TXRAs, can predict their activities, and can thus be used to design novel antagonists. Figure Optimal three-dimensional pharmacophore models of TXA₂ receptor antagonists (TXRAs) built with HypoGenRefine (a) and HipHop (b) modules. a Hypo-1 model mapped with compounds 11 (purple), and 20 (green). b Hypo-2 model mapped with compounds 31 (green) and 64 (yellow). Spheres: Green Hydrogen bond acceptors (HBA), cyanhydrophobic (H), orange aromatic rings (RA), black excluded volumes (EV) Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

烟草根际可培养微生物多样性及防病促生菌的筛选

【背景】根际微生物在植物根部生态系统中扮演着重要角色,影响着植物的营养吸收和健康生长。【目的】了解常年不发病烟田烤烟品种K326根际可培养微生物的多样性,筛选具有防病促生功能的菌株,为烟草病害绿色防控提供资源。【方法】采用传统培养方法对烟草根际土壤中的细菌和真菌进行分离鉴定,评价菌株的促生特性及病原菌拮抗能力,并进一步验证典型菌株对盆栽烟苗的促生效果。【结果】共获得261株微生物菌株,包括160株细菌和101株真菌。经分子鉴定,细菌中以变形菌门(Proteobacteria)和厚壁菌门(Firmicutes)为主要类群;真菌中以子囊菌门(Ascomycota)和毛霉菌门(Mucoromycota)为主要类群。在属水平上,细菌以假单胞菌属(Pseudomonas)和芽孢杆菌属(Bacillus)为主,真菌以曲霉属(Aspergillus)和青霉属(Penicillium)为主。从不同种水平上进一步选择44株细菌为代表菌株,发现它们均具有不同程度的吲哚-3-乙酸(Indole-3-Acetic Acid,IAA)产生能力,9株能够溶解有机磷,16株能够溶解无机磷,13株产生铁载体,14株产...     

Sexual identity differences in biomarkers of tobacco exposure among women in a national sample

BackgroundSexual minority women are consistently at increased risk for tobacco use compared to heterosexual women. Neither biomarkers of nicotine exposure nor biomarkers of tobacco toxicant exposure have been examined by sexual identity.MethodsThis study used interview and biomarker data from women in the biomarker core sample of Wave 1 of the Population Assessment of Tobacco and Health (PATH) study (2013–2014; n = 4930). We examined associations of sexual identity with nicotine exposure (measured with urinary cotinine and TNE-2) and with tobacco-specific nitrosamines (measured with urinary NNAL). Multivariable regression modeling was used to examine these associations among the full biomarker core sample, among past 30-day tobacco users, and among exclusive established cigarette users before and after controlling for tobacco use quantity and intensity.ResultsIn the full biomarker sample of women, prior to adjusting for tobacco use quantity and intensity, bisexual women had significantly higher cotinine, TNE-2, and NNAL levels compared to heterosexual women. Among exclusive established cigarette users, gay/lesbian women had significantly higher NNAL compared to heterosexual women prior to adjusting for tobacco quantity and intensity. No differences by sexual identity were found after adjusting for tobacco use quantity and intensity.ConclusionsThis is the first study to demonstrate differences in biological markers of tobacco exposure by sexual identity among women in the U.S. This has important public health implications as greater exposure to both nicotine and to tobacco-specific nitrosamines are strongly linked to cancer risk.     

Biodegradation of nicotine from tobacco waste extract by <Emphasis Type="Italic">Ochrobactrum intermedium</Emphasis> DN2

Ochrobactrum intermedium DN2 was used to degrade nicotine in tobacco waste extracts. The optimal temperature and pH of nicotine degradation by strain DN2 was 30–37 °C and 7.0, respectively. Under these optimal conditions, the average degradation rate of nicotine in a 30L fed-batch culture was 140.5 mg l⁻¹ h⁻¹. The results of this study indicate that strain DN2 may be useful for reducing the nicotine content of reconstituted tobacco.     

Structural insight into the inhibition of acetylcholinesterase by 2,3,4, 5-tetrahydro-1, 5-benzothiazepines

Benzothiazepines 1–3 inhibited acetylcholinesterase (AChE; EC 3.1.1.7) enzyme in a concentration-dependent fashion with IC₅₀ values of 1.0 ± 0.002, 1.2 ± 0.005 and 1.3 ± 0.001 μM, respectively. By using linear-regression equations, Lineweaver-Burk, Dixon plots and their secondary replots were constructed which indicated that compounds 1–3 are non-competitive inhibitors of AChE with K_i values of 0.8 ± 0.04, 1.1 ± 0.002, and 1.5 ± 0.001 μM, respectively. Molecular docking studies revealed that all the compounds are completely buried inside the aromatic gorge of AChE, extending deep into the gorge of AChE. A comparison of the docking results of compounds 1–3 displayed that these compounds generally adopt the same binding mode in the active site of AChE. The superposition of the docked structures demonstrated that the non-flexible benzothiazepine always penetrate into the aromatic gorge through the six-membered ring A, which allowed the ligands to interact simultaneously with more than one subsites of the active center of AChE. The higher AChE inhibitory potential of compounds 1–3 was found to be the cumulative effect of hydrophobic contacts and π-π interactions between the ligands and AChE. The relatively high affinity of benzothiazepine 1 with AChE was found to be due to additional hydrogen bond in benzothiazepine 1-AChE complex. The results indicated that substitution of halogen and methyl groups by hydrogen at aromatic ring of the benzothiazepine decreased the affinity of these molecules towards enzyme that may be due to the polar non-polar repulsions of these moieties with the amino acid residues in the active site of AChE. The observed binding modes of benzothiazepines 1–3 in the active site of AChE explain the affinities of benzothiazepines and provide a rational basis for the structure-based drug design of benzothiazepines with improved pharmacological properties.     

Cinnamic,myristic and fumaric acids in tobacco root exudates induce the infection of plants by Ralstonia solanacearum

Aim

The secretion of allelochemicals from plant roots plays a key role in soil sickness and soil-borne disease. The goal of this study was to investigate the role of allelopathic chemicals in Ralstonia solanacearum-infected tobacco roots.

Methods

The organic acids investigated in the present study are major components of tobacco root exudates. Through a swarming assay, we assessed the chemotaxis and colonization of R. solanacearum in response to organic acids.

Results

Fumaric acid was detected, and the results showed that this acid could serve as a semiochemical for attracting R. solanacearum and inducing the formation of biofilms of this species. The results also revealed that cinnamic and myristic acids play significant roles on swarming motility and chemotaxis. In addition, cinnamic, myristic and fumaric acids could enhance the expression of chemotaxis- and motility-related genes in R. solanacearum cultured in minimal medium. Furthermore, these three acids promote R. solanacearum colonization and accelerate disease progression in tobacco.

Conclusion

Cinnamic, myristic and fumaric acids could serve as semiochemical attractants to induce the colonization and infection of R. solanacearum. The results of the present study enhance our understanding of the ecological effects of plant root exudates in plant-microbe interactions and help to reveal the relationship between tobacco bacterial wilt and the autotoxins and allelochemicals that accumulate from root exudates.

     

烟草化感自毒物质降解复合菌剂的优化及应用效果评价

烟草(Nicotianatabacum)长期连作导致土壤中烟草自身分泌的化感自毒物质的积累,加剧了烟草青枯病(tobacco bacterial wilt, TBW)的发生,严重威胁烟草的生产并造成了巨大的经济损失。【目的】获得对长期连作烟田青枯病具有显著防治效果的复合菌剂。【方法】利用筛选到的烟草化感自毒物质降解菌株构建复合菌剂,通过正交试验和单因素试验对菌种配比和助剂进行优化,对其在温室盆栽和连作烟田生产的应用效果进行评价。【结果】正交试验结果表明,复合菌剂中芽孢杆菌(Bacillus sp.) NO1、布鲁氏菌(Brucella sp.) NO8、芽孢杆菌(Bacillus sp.) NO9和NO10的最佳菌种配比为1:3:4:2;单因素试验结果表明最佳的载体为白炭黑;最佳润湿剂和分散剂分别为六偏磷酸钠和丁基萘磺酸钠,其最佳比例为1:1,最佳用量各为2%;稳定剂甘油的最佳浓度为1.0%。盆栽试验结果显示,复合菌剂对6种化感自毒物质的降解率和对青枯病的抑制率均达到78%以上。大田试验结果显示,100倍稀释的复合菌剂对连作15年烟田中的6种化感自毒物质具有显著的降解效果,并且能明显增...     

一个产香大型真菌鉴定及其挥发性成分分析

【目的】开发优质的产香大型真菌资源,丰富挥发性香味成分的获取途径。【方法】本文通过传统形态分类学和分子生物学相结合的方法对采自泰国北部的野生产香大型真菌及其菌株进行鉴定。利用液体发酵、HP20大孔树脂固相萃取和气相色谱-质谱联用(GC-MS)等方法分析发酵液中的挥发性物质。同时筛选了对该真菌液体培养的最佳碳源、氮源及无机盐离子条件。【结果】该真菌经鉴定为茉莉小皮伞Marasmiusjasminodorus,分析发现其主要香味组分及对应的峰面积百分比分别为芳樟醇(33.11%)、5-羟甲基糠醛(4.64%)、4-甲基-5-(β羟乙基)噻唑(4.55%)、甲基麦芽酚(4.49%)、糠醇(4.46%)、桃醛(2.20%)、羟基丙酮(2.18%)等。同时该菌的最优培养液配方中最佳碳源、氮源及无机盐离子为麦芽糖、酵母粉和KH₂PO₄。【结论】本研究表明该大型真菌能够产生多种在现有工业生产中广泛应用的挥发性香味成分,如芳樟醇等,所以其在天然香精香料生产中具有较好的潜在应用前景。     

致病疫霉效应蛋白Pi16275的功能研究

[目的]分析致病疫霉效应蛋白Pi16275的超量表达对病原菌致病性的影响,明确Pi16275的亚细胞定位,筛选Pi16275在植物中的互作靶标蛋白及靶标蛋白在抵御病原菌侵染过程中的作用,初步揭示Pi16275在病原菌侵染植物过程中的作用机制.[方法]利用农杆菌介导的烟草瞬时表达系统在烟草叶片表皮细胞中瞬时表达Pi162...     

餐厨垃圾高温好氧生物减量菌种的筛选及特性

【背景】随着餐厨垃圾产生量的逐步提高,如何实现其快速降解,成为餐厨垃圾处理亟待解决的问题。餐厨垃圾的高温好氧生物减量技术是一种可以快速降解餐厨垃圾的有效方法。【目的】筛选能够适应餐厨垃圾环境且具有高效降解餐厨垃圾中有机物能力的菌株,以提高餐厨垃圾的降解效率和减量效果。【方法】采用温度梯度耐受性实验和餐厨垃圾浸出液高油高盐耐受性实验进行菌种初筛,并利用产酶培养基复筛及餐厨垃圾生物减量实验验证。【结果】通过初筛、复筛和功能验证,最终获得4株生物减量效果优良的菌株N3-1、C7、N3-3和G6-1,其对餐厨垃圾挥发性固体(volatile solid,VS)的降解率分别为36.95%、33.23%、32.83%和31.91%,是对照组的3.02、2.71、2.68和2.61倍。经鉴定,这4株菌分别属于热嗜油地芽孢杆菌(Geobacillus thermoleovorans)、史氏芽孢杆菌(Bacillus smithii)、热解木糖地芽孢杆菌(Geobacillus caldoxylosilyticus)和立陶宛地芽孢杆菌(Geobacillus lituanicus)。【结论】筛选出的4株菌均具有较强的餐厨垃圾原料适应性和高效的生物降解能力,为开发餐厨垃圾高温好氧复合菌剂奠定了基础,并为实现餐厨垃圾减量化、无害化处理和资源化利用提供了技术支持。     

酵母菌合成2-苯乙醇的研究进展

2-苯乙醇是一种具有令人愉悦的玫瑰风味的芳香醇,在食品、化妆品和药品等领域具有广泛的应用。本文对酵母菌合成2-苯乙醇的代谢途径及其调控过程、以及提高2-苯乙醇产量的国内外研究进展进行了综述,并对通过微生物转化法合成2-苯乙醇目前存在的不足及进一步研究方向进行了讨论。     

几株新鞘氨醇杆菌的趋化性及其相关基因组成特点

【目的】选择了几株隶属于新鞘氨醇杆菌属(Novosphingobium)的有和没有多环芳烃(PAHs)降解能力的细菌,在基因组水平比较分析它们的趋化通路,并探究其中一些菌株对芳香化合物和三羧酸(TCA)循环中间代谢产物的趋化性。【方法】通过基因组的比较分析,研究了几株新鞘氨醇杆菌的趋化蛋白组成和趋化基因分布,并采用滴定法和游动平板法检测了相关菌对芳香族化合物和TCA循环中间代谢产物的趋化性。【结果】N.pentaromativorans US6-1、N.pentaromativorans F2、Novosphingobium indicum K13、Novosphingobium stygium DSM 12445、Novosphingobium sp.C2AC等5株新鞘氨醇杆菌对芳香族化合物和TCA循环中间代谢物具有不同程度的趋化性;所选的7株已完成基因组测序的新鞘氨醇杆菌N.pentaromativorans F2、N.pentaromativorans US6-1、Novosphingobium sp.PP1Y、Novosphingobium sp.AP12、Novosphingobium sp.Rr 2-17,Novosphingobium sp.B-7和Novosphingobium sp.DSM 19370均含有趋化蛋白MCP、CheW、CheA、CheB、CheR和CheY,且亲缘关系最近的US6-1、F2和PP1Y 3株菌的che基因簇中的基因排列一致,为che W-Y-D-B-R-A-(X);新鞘氨醇杆菌的趋化系统属Fla类型。【结论】几株新鞘氨醇杆菌都具有较为完整的趋化通路,且对多种芳烃及其代谢物的趋化性各不相同,其中菌株US6-1趋化现象最明显。     

Analysis of 4-aminobiphenyl in smoker’s and nonsmoker’s urine by tandem mass spectrometry

The aromatic amine 4-aminobiphenyl (4-ABP) is present in tobacco smoke. In humans, it is also a known bladder carcinogen. We describe here a method for the quantification of total 4-ABP in urine using capillary gas chromatography/tandem mass spectrometry, with an effective detection limit in urine samples of approximately 0.87 pg/mL. We also examined the efficiency of chemical or enzymatic hydrolysis of urinary aromatic amine metabolites. Although we found acidic or basic hydrolysis effective, we found enzymatic hydrolysis (β-glucuronidase with either Escherichia coli or Helix pomatia) ineffective. As part of this work, we also confirm the presence of N-acetyl-4-ABP and 4-ABP glucuronide in human urine samples from smokers. These metabolites have been reported in animal studies, but previously they have not been identified in human samples. These metabolites, however, were found to be unstable and thus infeasible for biomonitoring. The final validated urinary total 4-ABP assay was applied to the analysis of samples from smokers and nonsmokers, whose status was confirmed from cotinine EIA measurements. Among 41 confirmed nonsmokers, the geometric mean (95% CI) of 4-ABP concentration was 1.64 pg/mg creatinine (1.30–2.07). Conversely, in 89 smokers, the geometric mean of 4-ABP concentration was significantly greater, at 8.69 pg/mg creatinine (7.43–10.16), p?<?0.001. Our results indicate that following tobacco smoke exposure, total urinary 4-ABP is a reliable biomarker for exposure to this carcinogen.     

Predicting bone turnover following tobacco exposure using bone alkaline phosphatase and N-telopeptide biomarkers and possible variability and effect modification of these markers by race/ethnicity

Abstract

Introduction: This study investigated the systemic response of serum bone alkaline phosphatase (SBAP) and urinary N-telopeptide (UNTX) to tobacco exposure and environmental tobacco smoke (ETS) and the possible effect modification (and variability) of this response by racial/ethnic origin.

Methods: Data (n?=?5411) were obtained from the National Health and Nutrition Examination Survey, with data analysis done on adults aged ≥ 20?years. Outcome variables were SBAP and UNTX. Independent variable was tobacco exposure measured using serum cotinine levels and adjusted for covariates. Generalized linear models were used to explore associations.

Results: A percentage increase in log transformed serum cotinine was associated with a 0.005 percentage increase in log transformed SBAP (CI: 0.002, 0.008) and 0.02 percentage increase in log transformed UNTX (CI: ?0.01, 0.04) with interaction between cotinine and race/ethnicity (p?=?0.01). Stratifying by race/ethnicity, tobacco exposure was associated with significant decreases in UNTX among non-Hispanic Whites – 0.008(?0.014, ?0.002) and Mexican Americans ?0.014 (?0.025, ?0.002) only. Categories of serum cotinine were associated with a monotonic increase in SBAP (p for trend <0.001) and monotonic non-linear decrease in UNTX (p for trend?>?0.05).

Conclusions: Tobacco and environmental tobacco exposure are associated with SBAP and increased bone formation. The response of UNTX to these exposures is modified by race/ethnicity with non-Hispanic Whites and Mexican-Americans less sensitive to the resorptive effects of tobacco exposure on bone.