The general procedure for the synthesis of CuS NPs in water was as follows. Into 1000 mL of aqueous solution of CuCl2 (0.1345 g, 1 mmol) and sodium citrate (0.2 g, 0.68 mmol) was added 1 mL of sodium sulfide solution (Na2S, 1 M) under stirring at room temperature. The pale blue CuCl2 solution turned dark brown immediately upon the addition of sodium sulfide. Five minutes later, the reaction mixture was heated to 90°C and stirred for 15 min until a dark green solution was obtained. The mixture was transferred to ice-cold water. The Cit-CuS NPs were obtained and stored at 4°C. To introduce PEG coating, about 1 mg of SH-PEG was added into the Cit-CuS NP solution (1.42×1015 NPs in 1.0 mL of water). The reaction was allowed to proceed overnight at room temperature.
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Sodium sulfide
Sodium sulfide
Sodium sulfide (Na2S) is an inorganic compound with a variety of industrial and scientific applications.
It is commonly used as a reducing agent, a precipitating agent, and in the production of other sulfur-containing compounds.
Sodium sulfide can be found in the form of hydrated crystals or as an aqueous solution.
It is an important chemical in various industrial processes, including leather tanning, pulp and paper production, and wastewater treatment.
Researchers studying sodium sulfide may be interested in exploring its synthesis, properties, and diverse applications through the use of PubCompare.ai's optimized research tools.
It is commonly used as a reducing agent, a precipitating agent, and in the production of other sulfur-containing compounds.
Sodium sulfide can be found in the form of hydrated crystals or as an aqueous solution.
It is an important chemical in various industrial processes, including leather tanning, pulp and paper production, and wastewater treatment.
Researchers studying sodium sulfide may be interested in exploring its synthesis, properties, and diverse applications through the use of PubCompare.ai's optimized research tools.
Most cited protocols related to «Sodium sulfide»
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The cytoarchitecture of the PFC was studied in ten adult, male mice (strain C57BL/6) of similar weight (approximately 20 g). These control mouse brains were kindly donated and immersion fixed by Dr. H. Manji, NIMH, USA. All animal procedures were in strict accordance with the NIH animal care guidelines. The histological processing of these brains was performed at the laboratory of Dr. Rajkowska. The brains were embedded in 12% celloidin, cut into 40-μm serial sections using a sliding microtome and Nissl (1% cresyl violet) stained. Celloidin was chosen as an embedding medium to allow for the preparation of ‘thick’ sections with clear morphology and high contrast of Nissl-stained neurons and glial cells. In these immersion-fixed brains, any spots showing pycnotic reaction were not incorporated in this study.
In addition to these ten mice, four adult male mice (C57BL/6 strain) were stained for dopamine and four adult male mice for AChE, myelin, and immunohistochemically for SMI, PV and CB. For each staining, a different set of sections with several consecutive sections stained with Nissl at HBMU’s laboratory was used. The antibodies applied were the dopamine (DA) antibody (Geffard et al. 1984 (link)), SMI-32 antibody (Sternberger Monoclonals Inc., Baltimore, MD, USA: monoclonal antibody to one epitope of non-phosphorylated tau neurofilaments, lot number 11), SMI-311antibody (pan-neuronal neurofilament marker cocktail of several monoclonal antibodies for several epitopes of non-phosphorylated tau protein, Sternberger Monoclonals Inc., Baltimore, MD, USA: lot number 9) (SMI antibodies are presently distributed through Covance Research Products, USA), monoclonal anti-CB D-28K antibody (Sigma, St. Louis, MO, USA: product number C-9848, clone number CB-955, lot number 015K4826), and monoclonal anti-PV antibody (Sigma, St. Louis, MO, USA: product number P-3171, clone number PA-235, lot number 026H4824). Mice to be stained for DA were intracardially perfused under deep pentobarbital anesthesia (1 ml/kg body weight, i.p.), with saline followed by fixative. For DA staining, the fixative was 5% glutaraldehyde in 0.05 M acetate buffer at pH 4.0. After perfusion, the brains were immersed in 0.05 Tris containing 1% sodium disulfite (Na2S2O5) at pH 7.2 (De Brabander et al. 1992 (link)). Mouse PFC was sectioned at 40 μm by a vibratome. These sections were stained overnight in a cold room at 4°C using the polyclonal primary antibody sensitive to DA that was raised in the Netherlands Institute for Brain Research (NIBR) (Geffard et al. 1984 (link)), the specificity of which had been demonstrated previously (Kalsbeek et al. 1990 (link)). DA antiserum was diluted 1:2,000 in 0.05 M Tris containing 1% Na2S2O5 and 0.5% Triton X-100, pH 7.2. After overnight incubation, the sections were washed three times with Tris-buffered saline (TBS) and subsequently incubated in the secondary antibody goat–antirabbit, also raised in NIBR at 1:100 for 1 h. After having been rinsed 3× in TBS, it was incubated in the tertiary antibody, peroxidase–antiperoxidase, at 1:1,000 for 60 min. Both the secondary and the tertiary antibodies were diluted in TBS with 0.5% gelatine and 0.5% Triton X-100. For visualization, the sections were transferred into 0.05% diaminobenzidine (DAB; Sigma) with 0.5% nickel ammonium sulfate. The reaction was stopped after a few minutes by transferring the sections to TBS (3 × 10 min), then the sections were mounted on slides, air dried, washed, dehydrated and coverslipped.
Mice to be stained with anti-PV, anti-CB and SMI-32 and SMI-311 were fixed with 4% formaldehyde solution in 0.1 M phosphate buffer at pH 7.6. Mouse PFC was sectioned at 40 μm by a vibratome. To prevent endogenous peroxidase activity, free-floating sections were pretreated for 30 min in a Tris-buffered saline (TBS) solution containing 3% hydrogen peroxide and 0.2% Triton X-100. To prevent non-specific antibody staining, these sections were placed in a milk solution (TBS containing 5% nonfat dry milk and 0.2% Triton X-100) for 1 h. Incubation of the primary antibody, directly after the milk step was carried out overnight in a cold room at 4°C. The primary antibodies were diluted in the above-mentioned milk solution: SMI-32 and SMI-311 at 1:1,000, PV antibody at 1:1,000, and CB antibody at 1:250. For the monoclonal SMI-32, SMI-311, PV and CB antibodies, raised in mice, we used peroxidase-conjugated rabbit–antimouse (1:100 in 5% milk solution with 0.2% Triton X-100) as a secondary antibody. Visualization took place in 0.05% diaminobenzidine enhanced with 0.2% nickel ammonium sulfate. The reaction was stopped after a few minutes by transferring these sections to TBS (3 × 10 min), after which the sections were rinsed in distilled water, mounted on slides, air dried, washed, dehydrated and coverslipped. Control sections that were incubated according to the same procedure as described above, omitting the primary antibody, were all negative. All sections were cut coronally, because the coronal plane offers in general the best view to differentiate between the subareas of the rodent PFC (Uylings et al. 2003 (link); Van de Werd and Uylings 2008 (link)).
Sections were processed for AChE staining according to the protocol described by Cavada et al. (1995 (link)). The sections were incubated overnight in a solution of cupric sulfate and acetate buffer at pH 5 to which acetylthiocholine iodide and ethopropazine were added just before the start of incubation. After rinsing, the sections were developed in a sodium sulfide solution until a light brown color appeared and subsequently intensified to a dark brown color in a silver nitrate solution. Finally, the sections were differentiated after rinsing in a thiosulfate solution, dehydrated and mounted. In all steps, the solutions and sections were shaken constantly. The myelin was stained with silver by physical development according to Gallyas (1979 (link)). The sections were first placed in 100% ethanol and then immersed in a 2:1 solution of pyridine and acetic acid for 30 min. After rinsing, they were placed in an ammonium silver nitrate solution and after rinsing with 0.5% acetic acid, the sections were immersed in the optimal physical developer solution at room temperature (Gallyas 1979 (link)) until they showed good stain intensity under the microscope. Then the development of the staining was stopped in 0.5% acetic acid and the sections were dehydrated and mounted with Histomount. The sections were studied at intervals of 80–160 μm, and examined under a light microscope at a 63× magnification.
In addition to these ten mice, four adult male mice (C57BL/6 strain) were stained for dopamine and four adult male mice for AChE, myelin, and immunohistochemically for SMI, PV and CB. For each staining, a different set of sections with several consecutive sections stained with Nissl at HBMU’s laboratory was used. The antibodies applied were the dopamine (DA) antibody (Geffard et al. 1984 (link)), SMI-32 antibody (Sternberger Monoclonals Inc., Baltimore, MD, USA: monoclonal antibody to one epitope of non-phosphorylated tau neurofilaments, lot number 11), SMI-311antibody (pan-neuronal neurofilament marker cocktail of several monoclonal antibodies for several epitopes of non-phosphorylated tau protein, Sternberger Monoclonals Inc., Baltimore, MD, USA: lot number 9) (SMI antibodies are presently distributed through Covance Research Products, USA), monoclonal anti-CB D-28K antibody (Sigma, St. Louis, MO, USA: product number C-9848, clone number CB-955, lot number 015K4826), and monoclonal anti-PV antibody (Sigma, St. Louis, MO, USA: product number P-3171, clone number PA-235, lot number 026H4824). Mice to be stained for DA were intracardially perfused under deep pentobarbital anesthesia (1 ml/kg body weight, i.p.), with saline followed by fixative. For DA staining, the fixative was 5% glutaraldehyde in 0.05 M acetate buffer at pH 4.0. After perfusion, the brains were immersed in 0.05 Tris containing 1% sodium disulfite (Na2S2O5) at pH 7.2 (De Brabander et al. 1992 (link)). Mouse PFC was sectioned at 40 μm by a vibratome. These sections were stained overnight in a cold room at 4°C using the polyclonal primary antibody sensitive to DA that was raised in the Netherlands Institute for Brain Research (NIBR) (Geffard et al. 1984 (link)), the specificity of which had been demonstrated previously (Kalsbeek et al. 1990 (link)). DA antiserum was diluted 1:2,000 in 0.05 M Tris containing 1% Na2S2O5 and 0.5% Triton X-100, pH 7.2. After overnight incubation, the sections were washed three times with Tris-buffered saline (TBS) and subsequently incubated in the secondary antibody goat–antirabbit, also raised in NIBR at 1:100 for 1 h. After having been rinsed 3× in TBS, it was incubated in the tertiary antibody, peroxidase–antiperoxidase, at 1:1,000 for 60 min. Both the secondary and the tertiary antibodies were diluted in TBS with 0.5% gelatine and 0.5% Triton X-100. For visualization, the sections were transferred into 0.05% diaminobenzidine (DAB; Sigma) with 0.5% nickel ammonium sulfate. The reaction was stopped after a few minutes by transferring the sections to TBS (3 × 10 min), then the sections were mounted on slides, air dried, washed, dehydrated and coverslipped.
Mice to be stained with anti-PV, anti-CB and SMI-32 and SMI-311 were fixed with 4% formaldehyde solution in 0.1 M phosphate buffer at pH 7.6. Mouse PFC was sectioned at 40 μm by a vibratome. To prevent endogenous peroxidase activity, free-floating sections were pretreated for 30 min in a Tris-buffered saline (TBS) solution containing 3% hydrogen peroxide and 0.2% Triton X-100. To prevent non-specific antibody staining, these sections were placed in a milk solution (TBS containing 5% nonfat dry milk and 0.2% Triton X-100) for 1 h. Incubation of the primary antibody, directly after the milk step was carried out overnight in a cold room at 4°C. The primary antibodies were diluted in the above-mentioned milk solution: SMI-32 and SMI-311 at 1:1,000, PV antibody at 1:1,000, and CB antibody at 1:250. For the monoclonal SMI-32, SMI-311, PV and CB antibodies, raised in mice, we used peroxidase-conjugated rabbit–antimouse (1:100 in 5% milk solution with 0.2% Triton X-100) as a secondary antibody. Visualization took place in 0.05% diaminobenzidine enhanced with 0.2% nickel ammonium sulfate. The reaction was stopped after a few minutes by transferring these sections to TBS (3 × 10 min), after which the sections were rinsed in distilled water, mounted on slides, air dried, washed, dehydrated and coverslipped. Control sections that were incubated according to the same procedure as described above, omitting the primary antibody, were all negative. All sections were cut coronally, because the coronal plane offers in general the best view to differentiate between the subareas of the rodent PFC (Uylings et al. 2003 (link); Van de Werd and Uylings 2008 (link)).
Sections were processed for AChE staining according to the protocol described by Cavada et al. (1995 (link)). The sections were incubated overnight in a solution of cupric sulfate and acetate buffer at pH 5 to which acetylthiocholine iodide and ethopropazine were added just before the start of incubation. After rinsing, the sections were developed in a sodium sulfide solution until a light brown color appeared and subsequently intensified to a dark brown color in a silver nitrate solution. Finally, the sections were differentiated after rinsing in a thiosulfate solution, dehydrated and mounted. In all steps, the solutions and sections were shaken constantly. The myelin was stained with silver by physical development according to Gallyas (1979 (link)). The sections were first placed in 100% ethanol and then immersed in a 2:1 solution of pyridine and acetic acid for 30 min. After rinsing, they were placed in an ammonium silver nitrate solution and after rinsing with 0.5% acetic acid, the sections were immersed in the optimal physical developer solution at room temperature (Gallyas 1979 (link)) until they showed good stain intensity under the microscope. Then the development of the staining was stopped in 0.5% acetic acid and the sections were dehydrated and mounted with Histomount. The sections were studied at intervals of 80–160 μm, and examined under a light microscope at a 63× magnification.
The mesophilic inoculum was taken from a biowaste co-utilizing wastewater treatment plant in Zirl (Austria) [25 (link)] with a reactor capacity of 1350 m3, an operation temperature of 39 (±0.2) °C, pH of 7.4 (±0.21), and total solids content of 2.2 (±0.04) g 100 g−1 fresh weight. The thermophilic inoculum was derived from the outlet sampling port of a thermophilic anaerobic digestion plant in Roppen (Austria) where about 2.500 tons of green waste and 6.200 tons of biowaste are treated per year [26 (link)], with a total reactor capacity of 900 m3, an operation temperature of 53 (±0.3) °C, pH of 7.9 (±0.44), and a total solids content of 26.2 (±2.0) g 100 g−1 fresh weight. Additional information regarding digester conditions and characteristics can be looked up elsewhere [22 (link)]. Plastic bottles filled with sludge were tightly sealed and immediately brought to the laboratory. For liquid handling, the sludge was sieved and diluted as described previously [27 (link),28 (link)]. The headspace was exchanged with a N2/CO2 (70:30)-gas mixture. The prepared samples were incubated at 37 °C and 52 °C for 15 days (mesophiles) and 20 days (thermophiles), respectively, until the sum of volatile fatty acids (VFA) was <200 mg kg−1. Subsequently, the samples were stored at 4 °C until further use.
Straw from grain (straw) was air-dried, but otherwise not chemically, physically, or biologically (pre)treated. The straw was cut into pieces 4–7 cm long. The C/N ratio of the straw (ratio: 56) was analysed with a TruSpec® CHN analyser (Leco, Germany) according to the manufacturer’s protocol. The straw was filled into 120 mL serum flasks, functioning as batch reactors, in different carbon-load concentrations with 3 (defined as low carbon load, LCL), 34 (defined as medium carbon load, MCL), and 170 (defined as high carbon load, HCL) mmol carbon-C reactor−1, respectively.
A basal anaerobic broth based on previous investigations [29 (link)] was prepared and modified as follows (per litre): 0.4 g NaCl, 0.4 g MgCl2 × 6 H2O, 0.68 g KH2PO4, 0.18 g NaOH, 0.05 g CaCl2 × 2 H2O, 0.4 g NH4Cl, 0.5 g L-cysteine, 10.0 g sodium carboxymethylcellulose (CMC), 0.5 g yeast extract, 2.0 g sodium acetate, 1.0 g sodium formiate, 1 mL vitamin solution [29 (link)], 1 mL trace element solution SL-10 (German Collection of Microorganisms and Cell Cultures GmbH (DSMZ), Braunschweig, Germany), 2 mL sodium sulfide solution (120 g L−1 Na2S), and 1 mL resazurine solution (1.15 g L−1 resazurine). After the pH was adjusted with 0.1 M sodium hydroxide to 7.5 ± 0.2, 48 mL of the medium was filled into the 120 mL serum flasks which had previously been filled with straw (as described above). A control containing the anaerobic broth but no straw was also included and equally treated thenceforward. The sealing and headspace gas exchange took place according to previous protocols [22 (link)]. The flasks were subsequently autoclaved and cooled down before further use.
For each temperature regime, a volume of 12 mL diluted inoculum was injected into each reactor. Subsequently, the reactors were incubated at 37 °C and 52 °C, respectively, extending over an anaerobic incubation period of 28 days. All variations were prepared in triplicate. Samples were taken on day 2, 4, 7, 14, 21, and 28. Liquid samples for pH, VFA, phenyl acids, and C/Nliquid were processed immediately or frozen at −20 °C. The pH of the samples was measured with pH indicator strips 4.0–9.0 (Merck, Germany).
For each temperature regime, a PCR-DGGE approach [30 (link),31 (link)] was conducted with all variants of day 0 to check for the same microbial community structure at the beginning of the experiment. Moreover, control samples of day 0, as well as samples of day 14 and 28 were used for next-generation sequencing (NGS) analyses.
VFA, total carbon, total nitrogen (C/Nliquid ratio), as well as phenyl acid analyses were done according to previous studies [22 (link),28 (link),32 (link)]. The gas over-pressure was measured with a GHM Greisinger GDH 200 sensor and used to calculate biogas and methane production [NmL] as described previously [27 (link)].
Liquid samples (1 mL) from day 0, 14, and 28 were centrifuged at 20,000 g for 15 min and resuspended in 1 mL sterile ¼ Ringer solution. Subsequently, DNA extraction was done using the Soil Extract II Kit DNA (Macherey-Nagel). 700 µL of each sample were filled in bead-tubes and centrifuged at 11,000 g for 10 min. The supernatant was discarded and buffer SL-1 (700 µL) and the enhanced lysis buffer (50 µL) were added. Each further extraction step was done according to the manufacturer’s manual. The DNA was eluted in 50 µL elution buffer. DNA quantity and co-extraction of contaminants (absorbance ratio 260/280 and 260/230) was checked via the NanoDrop 2000c™ system.
For the quantification of methanogenic Archaea, the mlas-f/mcrA-r primer pair [33 (link),34 (link)] targeting the methyl coenzyme M reductase subunit A (mcrA) gene was used. Analyses were done on a Corbett Life Science (Qiagen, the Netherlands) Rotor-Gene Q system. The PCR procedure was conducted as follows: initial denaturation at 95 °C for 10 min, followed by 45 cycles of denaturation (95 °C for 30 sec), annealing (66 °C for 30 sec), and extension (72 °C for 30 sec). A PCR solution of 20 µL contained 9 µL PCR Mix (SensiFast™ SYBR No-Rox Kit (2×) (Bioline, UK), 380 nM of each primer, 1 mM MgCl2, 20% Betaine Enhancer Solution (5×) (VWR International, Germany), and PCR-grade water to reach a final volume of 18 µL, as well as a 2 µL template (5 ng DNA µL−1). An eight-point standard curve using gene copies of Methanosarcina thermophila and a melt-curve analysis were included in the approach.
The NGS library was prepared in-house. The small subunit (SSU) rRNA gene primers 515f and 806r [35 (link)], according to the Earth Microbiome Project [36 (link)], were used to target the V4 region. The first PCR step, including the 16S rRNA primers and the Illumina® adapter sequences, was performed as follows: initial denaturation at 95 °C for 5 min, followed by 30 cycles of denaturation (95 °C for 45 s), annealing (57 °C for 45 s), and extension (72 °C for 90 s). A final extension step of 72 °C for 10 min was set at the end of the PCR process. A PCR solution of 25 µL contained 12 µL PCR Mix (VWR Red Taq DNA Polymerase Master Mix Kit (2×)), 250 nM of each primer-adapter combination, 20% Betaine Enhancer Solution (5×), PCR-grade water to reach final volume of 24 µL, as well as 1 µL DNA template (5 ng DNA µL−1). The quality of the PCR products was checked with a 1.5% agarose gel using the dye GelGreen® Nucleic Acid Gel Stain (Biotium, Fremont, CA, USA). The PCR products of the first step were diluted 1:5 and used as a template for a second amplification to attach the Illumina® barcodes (i5 and i7). The same PCR procedure as in the first PCR step was used, except that only seven cycles were applied and the annealing temperature was set to 56 °C. The PCR products were again checked with a 1.5% agarose gel. Subsequently, final PCR products were quantified fluorometrically, as described previously [37 (link)]. The PCR products (15 ng) of each sample were pooled and purified with a Hi Yield® Gel/PCR DNA Fragment Extraction Kit (SLG®, Gauting, Germany) and eluted in 50 µL Tris-HCl buffer. The DNA quantity was again measured via QuantiFluor® dsDNA Dye (Promega, Madison, WI, USA). Co-extraction of contaminants was checked via the NanoDrop 2000c™ system. The final ready-to-load sample pool showed a DNA concentration of 19 ng µL−1 (260/280 absorbance ratio: 1.88) and was subsequently sent to Microsynth AG in Switzerland where the sequencing was done according to the company’s protocols.
In total, 27 mesophilic, 27 thermophilic, as well as nine MOCK samples were analysed. Raw sample reads were processed using the program mothur version 1.39.5 [38 (link)] and the MiSeq SOP (July 2019) [39 (link)]. A contig file was created with the paired-end reads (4,428,969 sequences in total, 70,301 ± 14,082 sequences sample−1). After quality filtering (approx. 24% of the sequences were discarded), unique sequences were aligned to the SILVA V132 database (Appendix A ). After another quality check and pre-clustering, chimeric amplicons were removed applying the VSEARCH algorithm (VSEARCH v2.3.4.). Sequence classification was done with the k-nearest neighbor (knn) algorithm. Sequences were binned to phylotypes based on their taxonomy. For a better comparability of samples while simultaneously ensuring an adequate coverage of the species richness, rarefaction curves were generated, and samples were normalised to 22,800 reads per sample [40 (link)]. The Mantel test showed that the similarity matrices prior to and after rarefaction did not differ significantly from each other (R > 0.99, p < 0.01, N = 9999). Quality-filtered sequences were uploaded to GenBank® via the submission tool, BankIt (Appendix B ). Information on the MOCK communities can be looked up in Appendix C .
After quality filtering and subsampling to 22,800 reads per sample, a FASTA file containing only representative sequences and an operational taxonomic unit (OTU) table was generated via mothur (version 1.42.1). The files were uploaded tohttps://piphillin.secondgenome.com (September 2019). The tool piphillin used the nearest-neighbor algorithm to pair 16S rRNA gene sequences to genomes [41 (link)]. The Kyoto Encyclopedia of Genes and Genomes (KEGG) database [42 (link)] of October 2018 was applied. The identity cut-off was set at 97%. The analyses focused on general biochemical pathways and on pathways regarding anaerobic degradation/turnover of aromatic compounds: degradation of aromatic compounds (KEGG orthology ko01220), phenylpropanoid biosynthesis (KEGG orthology ko00940), benzoate degradation (KEGG orthology ko00362), and aminobenzoate degradation (KEGG orthology ko00627).
After rarefaction analyses, meso- and thermophilic data were analysed separately, using only OTUs with a total abundance of ≥35 for each temperature regime. In mothur, the get.coremicrobiome command was applied to gain information on the microorganisms being present in every variant of the respective temperature regime [38 (link),39 (link)]. For characterising microorganisms important for explaining the variation between the C-load samples (class) of each temperature regime (biomarker discovery), the LEfSe command was applied [43 (link)]. For an interactive visualisation of relative sequence abundances of meso- and thermophilic samples, respectively, the tool KRONA was used [44 (link)]. The significance cut-off was set at α = 0.05 for all analyses. Significant genera were shown with the program STAMP 2.1.3 (Parks et al., 2014). For that purpose, White’s non-parametric t-test (two-sided) was used to distinguish between variants [45 (link)]. Confidence intervals were provided via percentile bootstrapping (1000 permutations). The false discovery rate was controlled with the Benjamini-Hochberg procedure (B-H adjustment) [46 (link)]. Via the program PAST® 3 [47 ], Spearman’s rank correlation analyses (Spearman rs) were done for all samples of day 28 for each temperature regime: Genera with a standard deviation below 3 over all samples of day 28 of each temperature regime were excluded; phenyl acids were log (x+1), and the OTU data box-cox (x+1) transformed. The false discovery rate was controlled with the B-H adjustment in Microsoft® Excel®. Moreover, the Mantel test (Gower Similarity Index) was applied in PAST® 3. For piphillin and biochemical analyses, the Mann–Whitney U test (M-W, two-sided) and the Friedman ANOVA (time series) were applied, respectively (Statistica™ 13 (TIBCO® Software Inc.)). Graphical presentations of correlation analyses and methanogenic properties were done with SigmaPlot™ 14 (Systat® Software Inc.), of general microbial properties with STAMP 2.1.3, and of biochemical and piphillin analyses with Statistica™ 13.
Straw from grain (straw) was air-dried, but otherwise not chemically, physically, or biologically (pre)treated. The straw was cut into pieces 4–7 cm long. The C/N ratio of the straw (ratio: 56) was analysed with a TruSpec® CHN analyser (Leco, Germany) according to the manufacturer’s protocol. The straw was filled into 120 mL serum flasks, functioning as batch reactors, in different carbon-load concentrations with 3 (defined as low carbon load, LCL), 34 (defined as medium carbon load, MCL), and 170 (defined as high carbon load, HCL) mmol carbon-C reactor−1, respectively.
A basal anaerobic broth based on previous investigations [29 (link)] was prepared and modified as follows (per litre): 0.4 g NaCl, 0.4 g MgCl2 × 6 H2O, 0.68 g KH2PO4, 0.18 g NaOH, 0.05 g CaCl2 × 2 H2O, 0.4 g NH4Cl, 0.5 g L-cysteine, 10.0 g sodium carboxymethylcellulose (CMC), 0.5 g yeast extract, 2.0 g sodium acetate, 1.0 g sodium formiate, 1 mL vitamin solution [29 (link)], 1 mL trace element solution SL-10 (German Collection of Microorganisms and Cell Cultures GmbH (DSMZ), Braunschweig, Germany), 2 mL sodium sulfide solution (120 g L−1 Na2S), and 1 mL resazurine solution (1.15 g L−1 resazurine). After the pH was adjusted with 0.1 M sodium hydroxide to 7.5 ± 0.2, 48 mL of the medium was filled into the 120 mL serum flasks which had previously been filled with straw (as described above). A control containing the anaerobic broth but no straw was also included and equally treated thenceforward. The sealing and headspace gas exchange took place according to previous protocols [22 (link)]. The flasks were subsequently autoclaved and cooled down before further use.
For each temperature regime, a volume of 12 mL diluted inoculum was injected into each reactor. Subsequently, the reactors were incubated at 37 °C and 52 °C, respectively, extending over an anaerobic incubation period of 28 days. All variations were prepared in triplicate. Samples were taken on day 2, 4, 7, 14, 21, and 28. Liquid samples for pH, VFA, phenyl acids, and C/Nliquid were processed immediately or frozen at −20 °C. The pH of the samples was measured with pH indicator strips 4.0–9.0 (Merck, Germany).
For each temperature regime, a PCR-DGGE approach [30 (link),31 (link)] was conducted with all variants of day 0 to check for the same microbial community structure at the beginning of the experiment. Moreover, control samples of day 0, as well as samples of day 14 and 28 were used for next-generation sequencing (NGS) analyses.
VFA, total carbon, total nitrogen (C/Nliquid ratio), as well as phenyl acid analyses were done according to previous studies [22 (link),28 (link),32 (link)]. The gas over-pressure was measured with a GHM Greisinger GDH 200 sensor and used to calculate biogas and methane production [NmL] as described previously [27 (link)].
Liquid samples (1 mL) from day 0, 14, and 28 were centrifuged at 20,000 g for 15 min and resuspended in 1 mL sterile ¼ Ringer solution. Subsequently, DNA extraction was done using the Soil Extract II Kit DNA (Macherey-Nagel). 700 µL of each sample were filled in bead-tubes and centrifuged at 11,000 g for 10 min. The supernatant was discarded and buffer SL-1 (700 µL) and the enhanced lysis buffer (50 µL) were added. Each further extraction step was done according to the manufacturer’s manual. The DNA was eluted in 50 µL elution buffer. DNA quantity and co-extraction of contaminants (absorbance ratio 260/280 and 260/230) was checked via the NanoDrop 2000c™ system.
For the quantification of methanogenic Archaea, the mlas-f/mcrA-r primer pair [33 (link),34 (link)] targeting the methyl coenzyme M reductase subunit A (mcrA) gene was used. Analyses were done on a Corbett Life Science (Qiagen, the Netherlands) Rotor-Gene Q system. The PCR procedure was conducted as follows: initial denaturation at 95 °C for 10 min, followed by 45 cycles of denaturation (95 °C for 30 sec), annealing (66 °C for 30 sec), and extension (72 °C for 30 sec). A PCR solution of 20 µL contained 9 µL PCR Mix (SensiFast™ SYBR No-Rox Kit (2×) (Bioline, UK), 380 nM of each primer, 1 mM MgCl2, 20% Betaine Enhancer Solution (5×) (VWR International, Germany), and PCR-grade water to reach a final volume of 18 µL, as well as a 2 µL template (5 ng DNA µL−1). An eight-point standard curve using gene copies of Methanosarcina thermophila and a melt-curve analysis were included in the approach.
The NGS library was prepared in-house. The small subunit (SSU) rRNA gene primers 515f and 806r [35 (link)], according to the Earth Microbiome Project [36 (link)], were used to target the V4 region. The first PCR step, including the 16S rRNA primers and the Illumina® adapter sequences, was performed as follows: initial denaturation at 95 °C for 5 min, followed by 30 cycles of denaturation (95 °C for 45 s), annealing (57 °C for 45 s), and extension (72 °C for 90 s). A final extension step of 72 °C for 10 min was set at the end of the PCR process. A PCR solution of 25 µL contained 12 µL PCR Mix (VWR Red Taq DNA Polymerase Master Mix Kit (2×)), 250 nM of each primer-adapter combination, 20% Betaine Enhancer Solution (5×), PCR-grade water to reach final volume of 24 µL, as well as 1 µL DNA template (5 ng DNA µL−1). The quality of the PCR products was checked with a 1.5% agarose gel using the dye GelGreen® Nucleic Acid Gel Stain (Biotium, Fremont, CA, USA). The PCR products of the first step were diluted 1:5 and used as a template for a second amplification to attach the Illumina® barcodes (i5 and i7). The same PCR procedure as in the first PCR step was used, except that only seven cycles were applied and the annealing temperature was set to 56 °C. The PCR products were again checked with a 1.5% agarose gel. Subsequently, final PCR products were quantified fluorometrically, as described previously [37 (link)]. The PCR products (15 ng) of each sample were pooled and purified with a Hi Yield® Gel/PCR DNA Fragment Extraction Kit (SLG®, Gauting, Germany) and eluted in 50 µL Tris-HCl buffer. The DNA quantity was again measured via QuantiFluor® dsDNA Dye (Promega, Madison, WI, USA). Co-extraction of contaminants was checked via the NanoDrop 2000c™ system. The final ready-to-load sample pool showed a DNA concentration of 19 ng µL−1 (260/280 absorbance ratio: 1.88) and was subsequently sent to Microsynth AG in Switzerland where the sequencing was done according to the company’s protocols.
In total, 27 mesophilic, 27 thermophilic, as well as nine MOCK samples were analysed. Raw sample reads were processed using the program mothur version 1.39.5 [38 (link)] and the MiSeq SOP (July 2019) [39 (link)]. A contig file was created with the paired-end reads (4,428,969 sequences in total, 70,301 ± 14,082 sequences sample−1). After quality filtering (approx. 24% of the sequences were discarded), unique sequences were aligned to the SILVA V132 database (
After quality filtering and subsampling to 22,800 reads per sample, a FASTA file containing only representative sequences and an operational taxonomic unit (OTU) table was generated via mothur (version 1.42.1). The files were uploaded to
After rarefaction analyses, meso- and thermophilic data were analysed separately, using only OTUs with a total abundance of ≥35 for each temperature regime. In mothur, the get.coremicrobiome command was applied to gain information on the microorganisms being present in every variant of the respective temperature regime [38 (link),39 (link)]. For characterising microorganisms important for explaining the variation between the C-load samples (class) of each temperature regime (biomarker discovery), the LEfSe command was applied [43 (link)]. For an interactive visualisation of relative sequence abundances of meso- and thermophilic samples, respectively, the tool KRONA was used [44 (link)]. The significance cut-off was set at α = 0.05 for all analyses. Significant genera were shown with the program STAMP 2.1.3 (Parks et al., 2014). For that purpose, White’s non-parametric t-test (two-sided) was used to distinguish between variants [45 (link)]. Confidence intervals were provided via percentile bootstrapping (1000 permutations). The false discovery rate was controlled with the Benjamini-Hochberg procedure (B-H adjustment) [46 (link)]. Via the program PAST® 3 [47 ], Spearman’s rank correlation analyses (Spearman rs) were done for all samples of day 28 for each temperature regime: Genera with a standard deviation below 3 over all samples of day 28 of each temperature regime were excluded; phenyl acids were log (x+1), and the OTU data box-cox (x+1) transformed. The false discovery rate was controlled with the B-H adjustment in Microsoft® Excel®. Moreover, the Mantel test (Gower Similarity Index) was applied in PAST® 3. For piphillin and biochemical analyses, the Mann–Whitney U test (M-W, two-sided) and the Friedman ANOVA (time series) were applied, respectively (Statistica™ 13 (TIBCO® Software Inc.)). Graphical presentations of correlation analyses and methanogenic properties were done with SigmaPlot™ 14 (Systat® Software Inc.), of general microbial properties with STAMP 2.1.3, and of biochemical and piphillin analyses with Statistica™ 13.
2-acetylpyridine
Acetate
acetonitrile
Anabolism
Bicarbonate, Sodium
bis(tert-butoxycarbonyl)oxide
Boranes
Carbodiimides
Carbonates
Celite
hydrazine hydrate
indole
Iris Plant
lithium hydroxide monohydrate
Methyl Chloride
methyl iodide
Oxides
Palladium
Phosphorus
Silver
sodium hydride
Sulfides
triphenylphosphine
Most recents protocols related to «Sodium sulfide»
Each recipient was assigned to receive either saline control or one of the two experimental compounds, sodium iodide or hydrogen sulfide:
Saline (control): treatment of the flap with 0.9% saline solution.
Sodium iodide (NaI): solution containing 10 mg/mL. Animals in this group received an additional intravenous dose of 1 mg/mL one minute after the anastomosis was unclamped.
Hydrogen sulfide (H2S): solution containing 2 mg/kg.
To assess the effect of CO on the respiration of wild-type E. coli cells (strain MG1655), we investigated aerobic cultures in which a change in oxidase expression from cytochrome bo3 to the bd-type cytochromes is expected to occur during cell growth, following a progressive reduction in the availability of O2 in the medium, taking into account the striking difference between the oxidases in sensitivity to sodium sulfide, according to Forte et al. [77 (link)]. When cells grown in Luria–Bertani (LB) medium are assayed in an early growth phase of the culture (OD600 < 0.8), most of the respiration (50–70%) is sensitive to 50 μM sodium sulfide, indicating a prevalent expression of cytochrome bo3. In contrast, in a late growth phase of the culture (OD600 > 2.5), when oxygen is limiting, sodium sulfide causes only marginal effects on respiration, indicating a prevalent expression of the bd-type cytochromes.
Mice were deeply anesthetized with isoflurane and perfused with a 0.3% sodium sulfide solution, followed by saline and 4% paraformaldehyde (PFA). The brains were isolated and fixed with 4% PFA, and then dehydrated with 30% sucrose. The fresh-frozen tissue was sectioned using cryostat (Leica CM1950, Germany) at 25 μm thickness. These slices were placed in a developing solution at 26°C, and then immersed in a 5% sodium metabisulfite solution for 10 min to halt the metal self-developing reaction. Following dehydration and permeabilization, the slices were placed on glass microscope slides, sealed with neutral tree gum and observed using an optical microscope (Olympus BX63, Japan). The developer solution was prepared according to sodium sulfide Timm's method (Danscher, 1996 (link)).
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Ethylenediaminetetraacetate (EDTA), (country of manufacture China). Potassium permanganate (KMnO4), (country of manufacture China). Sodium sulfide (Na2S), (country of manufacture China). Water (H2O), (country of manufacture China).
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Sodium sulfide is a chemical compound with the formula Na2S. It is a white to yellowish crystalline solid that is soluble in water. Sodium sulfide is commonly used as a reducing agent, a precipitating agent, and a raw material in the production of other chemicals.
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Sodium sulfide nonahydrate is a chemical compound with the formula Na2S·9H2O. It is a crystalline solid that is commonly used as a reducing agent and a source of sulfide ions in various industrial and laboratory applications.
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Sodium hydroxide is a chemical compound with the formula NaOH. It is a white, odorless, crystalline solid that is highly soluble in water and is a strong base. It is commonly used in various laboratory applications as a reagent.
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Methanol is a clear, colorless, and flammable liquid that is widely used in various industrial and laboratory applications. It serves as a solvent, fuel, and chemical intermediate. Methanol has a simple chemical formula of CH3OH and a boiling point of 64.7°C. It is a versatile compound that is widely used in the production of other chemicals, as well as in the fuel industry.
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Ethanol is a clear, colorless liquid chemical compound commonly used in laboratory settings. It is a key component in various scientific applications, serving as a solvent, disinfectant, and fuel source. Ethanol has a molecular formula of C2H6O and a range of industrial and research uses.
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Hydrochloric acid is a commonly used laboratory reagent. It is a clear, colorless, and highly corrosive liquid with a pungent odor. Hydrochloric acid is an aqueous solution of hydrogen chloride gas.
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Silver nitrate is a chemical compound with the formula AgNO3. It is a colorless, water-soluble salt that is used in various laboratory applications.
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Sodium sulfide is a chemical compound used in various industrial and laboratory applications. It is a white to yellow crystalline solid with the chemical formula Na2S. Sodium sulfide is soluble in water and is commonly used as a reducing agent, a sulfur source, and a precipitating agent in various chemical processes.
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Acetone is a colorless, volatile, and flammable liquid. It is a common solvent used in various industrial and laboratory applications. Acetone has a high solvency power, making it useful for dissolving a wide range of organic compounds.
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NaCl is a chemical compound commonly known as sodium chloride. It is a white, crystalline solid that is widely used in various industries, including pharmaceutical and laboratory settings. NaCl's core function is to serve as a basic, inorganic salt that can be used for a variety of applications in the lab environment.
More about "Sodium sulfide"
Sodium sulfide (Na2S) is a versatile inorganic compound with a wide range of industrial and scientific applications.
It is commonly referred to as sodium sulfide or sodium monosulfide.
This chemical is frequently used as a reducing agent, precipitating agent, and in the production of other sulfur-containing compounds.
Sodium sulfide can be found in the form of hydrated crystals or as an aqueous solution.
Sodium sulfide is an important chemical in various industrial processes, such as leather tanning, pulp and paper production, and wastewater treatment.
Researchers studying this compound may be interested in exploring its synthesis, properties, and diverse applications.
For example, sodium sulfide nonahydrate (Na2S·9H2O) is a common hydrated form of the compound.
In addition to its industrial uses, sodium sulfide can also be utilized in chemical reactions involving other compounds like sodium hydroxide, methanol, ethanol, hydrochloric acid, silver nitrate, and acetone.
These reactions can lead to the formation of sodium chloride (NaCl) and other products.
PubCompare.ai's optimized research tools can help researchers efficiently explore the literature, preprints, and patents related to sodium sulfide, allowing them to identify the most accurate and effective experimental methods for their studies.
It is commonly referred to as sodium sulfide or sodium monosulfide.
This chemical is frequently used as a reducing agent, precipitating agent, and in the production of other sulfur-containing compounds.
Sodium sulfide can be found in the form of hydrated crystals or as an aqueous solution.
Sodium sulfide is an important chemical in various industrial processes, such as leather tanning, pulp and paper production, and wastewater treatment.
Researchers studying this compound may be interested in exploring its synthesis, properties, and diverse applications.
For example, sodium sulfide nonahydrate (Na2S·9H2O) is a common hydrated form of the compound.
In addition to its industrial uses, sodium sulfide can also be utilized in chemical reactions involving other compounds like sodium hydroxide, methanol, ethanol, hydrochloric acid, silver nitrate, and acetone.
These reactions can lead to the formation of sodium chloride (NaCl) and other products.
PubCompare.ai's optimized research tools can help researchers efficiently explore the literature, preprints, and patents related to sodium sulfide, allowing them to identify the most accurate and effective experimental methods for their studies.