The concentration of thiol collectors (PEX, SDD, IET and DDA) was determined by a UV–vis spectroscopic method [10 (link),11 (link),13 (link),30 (link)]. The absorbance of collector solution was recorded by a UV–vis spectrophotometer (UV-5500PC, Shanghai Metash Instruments Co. Ltd, China). The COD was determined by the standard dichromate method (HJ/T 399-2007). The TOC of water samples was measured by using a Shimadzu TOC-V organic carbon analyzer. The carbon mineralization extent (γc) of the collector (PEX, SDD, IET and DDA) was calculated as the below equation where TOC0 and TOCt (mg l−1) were the TOC concentration at initial and time t, respectively.
Uv 5500pc
Manufactured by Metash
Sourced in China
The UV-5500PC is a spectrophotometer that uses ultraviolet and visible light to analyze the absorption or transmission properties of samples. It is capable of measuring the absorbance or transmittance of light within the wavelength range of 190 to 1100 nanometers.
Automatically generated - may contain errors
Lab products found in correlation
Toc 5 organic carbon analyzer, by Shimadzu (3 mentions)
Precellys 24 homogenizer, by Bertin Technologies (1 mentions)
Ros assay kit, by Beyotime (1 mentions)
Apoptosis detection kit, by Beyotime (1 mentions)
Fetal bovine serum (fbs), by ExCell Bio (1 mentions)
400 mhz nmr spectrometer, by Bruker (1 mentions)
Zebron zb ffap, by Phenomenex (1 mentions)
Gcms qp2010 se, by Shimadzu (1 mentions)
15 protocols using uv 5500pc
1
Determination of Thiol Collector Concentration
2
Quantification of Intracellular Glutathione
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Streptococcus thermophilus was cultured to the stationary phase and collected by centrifugation (10,000 × g, 10 min). The cells were washed twice with 0.85% sodium chloride solution, and then resuspended with 1 mL of phosphate buffer B (0.2 M Na 2 HPO 4 , 0.2 M NaH 2 PO 4 , 2 mM EDTA, pH 7.0, stored at 4°C after sterilization). We added 0.1 g of glass beads into 500 μL of the bacterial solution. The mixture was shaken 3 times (6,000 × g, 30 s, 4°C) by a Precellys24 homogenizer (Bertin). The cell debris was discarded by centrifugation (12,000 × g, 10 min, 4°C), and the supernatant was collected to detect the GSH content.
The 35 μL of 0.3 mM NADPH, 10 μL of GSH sample, and 5 μL of 6 mM 5,5-dithiobis (2-nitrobenzoic acid; DTNB) was made up a 50 μL reaction system and equilibrated at 25°C. Subsequently, 1 μL of 50 U/mL of GSH reductase (Sangon Bioengineering Co., Ltd.) from the baker's yeast was added to the pre-warmed solution and transferred to a cuvette with a 1-cm light path. Finally, the samples were continuously monitored for changes in absorbance at a wavelength of 412 nm using a spectrophotometer (UV-5500PC; Shanghai Metash Instruments Co., Ltd.). We prepared NADPH, DTNB, GSH, and GSH reductase all in a phosphate buffer (0.125 M K 3 PO 4 , 6.3 mM EDTA, pH 7.5).
The 35 μL of 0.3 mM NADPH, 10 μL of GSH sample, and 5 μL of 6 mM 5,5-dithiobis (2-nitrobenzoic acid; DTNB) was made up a 50 μL reaction system and equilibrated at 25°C. Subsequently, 1 μL of 50 U/mL of GSH reductase (Sangon Bioengineering Co., Ltd.) from the baker's yeast was added to the pre-warmed solution and transferred to a cuvette with a 1-cm light path. Finally, the samples were continuously monitored for changes in absorbance at a wavelength of 412 nm using a spectrophotometer (UV-5500PC; Shanghai Metash Instruments Co., Ltd.). We prepared NADPH, DTNB, GSH, and GSH reductase all in a phosphate buffer (0.125 M K 3 PO 4 , 6.3 mM EDTA, pH 7.5).
3
Colorimetric Determination of Total Flavonoids
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The content of total flavonoid in FSI was measured by a colorimetric determination according to Heimler, D, et al. [40 (link)] with slight modifications in the study. One milliliter of the suitably diluted sample, 5 mL deionized water, and 1 mL 5% NaNO2 solution were added, incubated for 6 min; then 1 mL freshly prepared 10% Al(NO3)3 solution was mixed, incubated for 6 min; finally 10 mL of 1 mol/L sodium hydroxide solution was added. The final volume was added to 25 mL with deionized water. The mixture was kept at room temperature for 15 min, after which its absorbance value was read at 511 nm with an ultraviolet spectrophotometer (UV−5500PC, METASH, Shanghai, China) against the mixture without the sample. The content of total flavonoids was calculated by the calibration curve of rutin (rutin g/g sample). The regression data was listed in Table 3 .
4
Quantifying Lichen Phosphomonoesterase Activity
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Phosphomonoesterase activity of lichen thalli was determined as the method of Turner et al. (2001) (link). Briefly, in each nitrogen treatment, three lichen individuals were randomly selected from test 1.2.2, a piece of thallus of 2 cm length was cut from each thalli, then was added to 2.9 mL of 0.02 M citric acid–trisodium citrate solution. Then, 0.1 ml para-nitrophenylphosphate (pNPP) was added to the solution to reach a final concentration of 3 mM. Samples were subsequently incubated for 20 min at 15°C in a shaking water-bath in the dark. The reaction was terminated by transferring 2.5 ml of the essay solution into 0.25 ml terminator solution (1.1 M NaOH, 27.5 mM EDTA and 0.55 M K2HPO4). The absorbance was measured at a wavelength of 405 nm using a spectrophotometer (UV-5500PC, Metash instruments Co., Ltd., China). Thalli were then oven dried for 24 h at 65°C and weighed. Enzyme activity was expressed as μmol substrate hydrolysed g-1 dry mass h-1 using p-nitrophenol to calibrate the assay. Three replicates were performed for each nitrogen treatment.
5
Lichen Glutamine Synthetase Activity
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Six groups of lichens were incubated 2 months under a condition, same to the steps described in Section “Adsorption of Ammonium and Nitrate.” We sprayed lichens every 3 days with one of 0.1, 0.2, 0.4, 0.8, 1.6, and 2.0 mM NH4NO3 solution. Glutamine synthetase activity (GSA) was measured as Zhao et al. (2002) . In detail, 50 mg air dried lichen was manually grounded with 2 ml 5 mM phosphoric acid buffer (containing 0.5 mM EDTA, 50 mM K2SO4, pH 7.2), then the buffer was centrifuged 20 min (18000 rpm, 4°C). Subsequently, 1.2 ml crude enzyme liquid, 0.6 ml imidazole-HCl buffer (0.25 M), 0.4 ml glutamic sodium (0.30 M),0.4 ml ATP-Na (30 mM), 0.2 ml MgSO4 (0.5 M) were added into a tube. The tube was incubated in a 25°C water bath for 5 min, then 0.2 ml hydroxylamine (0.5 M) was added in. Fifteen minutes later, 0.8 ml ferric trichloride solution was added to terminate the reaction. We centrifuged the reaction solution 15 min (4500 rpm, 4°C), and absorption value (A) of the supernate was measured at 540 nm with spectrophotometer (UV-5500PC, Metash instruments Co., Ltd., China). Relative GSA was presented as A⋅h-1⋅g-1⋅DW, in which A was the absorption value of supernate. Three replicates were performed for each nitrogen treatment.
6
Characterization and Stability of PSSP-Coated ART-ISMN Nanoparticles
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ART and ISMN were purchased from Energy Chemical (Shanghai, China), mPEG5000 was sourced from AVT Pharmaceutical Tech Co., Ltd (Shanghai, China), apoptosis detection kit, ROS assay kit, and NO fluorescence probe were obtained from Beyotime (Shanghai, China), and RNS fluorescence probe was provided by BestBio (Nanjing, China). The polyclonal antibody was bought from Bioss (Beijing, China). Cell culture vessels were purchased from Nest Biotechnology (Wuxi, China).
1H NMR spectra were recorded on a 400 MHz NMR spectrometer (Bruker, Switzerland). NP size and zeta potential were obtained via dynamic light scattering (DLS) analysis (Malvern, UK). The morphology and diameter were characterized using a transmission electron microscope (TEM). Drug loading rates for different NP formulations were measured using the ultraviolet (UV)-vis spectrum (Metash, UV-5500PC, China). The stability of PSSP@ART-ISMN under physiological conditions was investigated in 10% fetal bovine serum (Excell Bio, China) over the course of seven days using DLS.
1H NMR spectra were recorded on a 400 MHz NMR spectrometer (Bruker, Switzerland). NP size and zeta potential were obtained via dynamic light scattering (DLS) analysis (Malvern, UK). The morphology and diameter were characterized using a transmission electron microscope (TEM). Drug loading rates for different NP formulations were measured using the ultraviolet (UV)-vis spectrum (Metash, UV-5500PC, China). The stability of PSSP@ART-ISMN under physiological conditions was investigated in 10% fetal bovine serum (Excell Bio, China) over the course of seven days using DLS.
7
Nitrate Reductase Activity in Lichens
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Nitrate reductase activity (NRA) was determined as the methods of Zhao et al. (2002) . About 50 mg lichen apex was submerged in to a tube containing 10 ml solution of 0.10 mol⋅L-1 KNO3. Then the tube was vacuumized 30 min, subsequently N2 was supplied into the tubes. Thereafter, tube was incubated in a dark, 25°C chamber for 30 min. Then enzyme reaction was terminated by taking the lichen apex out of the tube. Absorption of reaction solution was measured with spectrophotometer at 540 nm (UV-5500PC, Metash instruments Co., Ltd., China), as well dry mass of the lichen apex was measured. NRA was expressed as mg NO2-⋅h-1⋅g-1⋅DW. Each nitrogen treatment was replicated three times.
8
Quantifying Flavonoids in Fractionated Seaweed Extract
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Total flavonoid contents of FSR were measured using the aluminum chloride colorimetric assay49 with slight modification. Diluted FSR (concentration at 1.6%, v/v) was used in the analysis. A 40-μl aliquot of FSR was added into a 5-ml test tube containing 1.31 ml of distilled water and 75 μl of 5% (w/v) NaNO2 was then added. After 5 min, 75 μl 10% (w/v) AlCl3 was added and allowed to react for 6 min before 1 ml of 4% (w/v) NaOH dissolved in distilled water was added. The solution was mixed and kept for 12 min at room temperature before ultraviolet absorbance was measured against the control at 510 nm using a UV-5500PC (METASH). Control samples contained the same amount of chemicals except the FSR was replaced with distilled water. Rutin was used as a standard for constructing a calibration curve. Total flavonoid contents were expressed as mg rutin equivalents per of FSR.
9
Quantifying Total Phenolic Content in FSR
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Total phenolic contents of FSR were determined spectrophotometrically using Folin–Ciocalteu’s method as described by Kang et al.48 with slight modification. Diluted FSR (0.4%, v/v) was used in the analysis. A 10-μl aliquot of FSR was mixed with 0.25 ml 1 N Folin–Ciocalteu’s reagent in a 5-ml test tube. The mixture was covered and kept still for 2 min in the dark before 0.5 ml 12% (w/v) aqueous solution of Na2CO3 and 1.24 ml distilled water were added. The mixture was incubated for 1 h at room temperature and then ultraviolet absorbance was measured at 765 nm using a UV-5500PC (METASH) against a control. The control sample contained the same amount of chemicals with the FSR replaced by distilled water. Gallic acid (GA) was used as a standard for preparing a calibration curve. Total phenolic contents were expressed as mg GA equivalents per FSR.
10
Tyrosinase Inhibitory Activity Assay
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Tyrosinase inhibitory activity was examined in vitro as described by Elena et al.50 (link). Different concentrations of FSR or kojic acid (KA) were mixed with tyrosinase dissolved in 0.2 M phosphate buffer (pH 6.8) and kept still for 15 min at 37 °C. Next, 12.5 mM L-Dopa dissolved in 0.2 M phosphate buffer (pH 6.8) was added, so that the final concentration of tyrosinase was 25 U/ml and that of L-Dopa was 1.25 mM in the solution. Absorbance at 475 nm was measured using a UV-5500PC (METASH) after incubation for 5 min at room temperature. FSR was replaced by the equivalent amount of phosphate buffer in the control mixture prepared following the same procedures. The percentage of inhibition was calculated using the following equation: ODcontrol is the absorbance of the negative control and ODsample is the absorbance of the sample. KA served as a positive control. IC50 values were calculated as the concentration of causing a 50% inhibition of tyrosinase.
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