Acrylate terminated polymers were sampled from reaction vials prior to end-capping reactions and precipitated twice in 10x volumes of diethyl ether to recover neat polymer. Acrylate terminated polymers were then dried under vacuum for 2h and analyzed via 1H NMR in CDCl3 (Bruker 500 MHz) to confirm the presence of acrylate peaks and quantify degree of branching. End-capped polymer likewise was characterized via 1H NMR in CDCl3 to confirm complete reaction of end-cap monomer with acrylate terminated polymers. End-capped polymer was also characterized via gel permeation chromatography (GPC) using a Waters system with autosampler, styragel column and refractive index detector to determine MN, MW and polydispersity index (PDI) relative to linear polystyrene standards. GPC measurements were performed as previously described with minor changes of flow rate (0.5 mL/min) and increase in sample run time to 75 minutes per sample.32 (link)
Refractive index detector
Manufactured by Waters Corporation
Sourced in United States, Japan
The refractive index detector is a laboratory instrument used to measure the refractive index of a sample. It operates by passing a beam of light through the sample and measuring the change in the angle of the light as it passes through the sample. The refractive index is a fundamental property of a material and is used in various applications, such as chemical analysis and material characterization.
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Lab products found in correlation
Xbridge c8 column, by Waters Corporation (2 mentions)
500 mhz nmr, by Bruker (2 mentions)
Hplc system, by Waters Corporation (2 mentions)
Hpx 87h column, by Bio-Rad (2 mentions)
Styragel column, by Waters Corporation (1 mentions)
Lcms 1290 6460, by Waters Corporation (1 mentions)
Hlc 8320gpc ecosec, by Tosoh (1 mentions)
Sugar pak 1 column, by Waters Corporation (1 mentions)
High performance liquid chromatography (hplc), by Waters Corporation (1 mentions)
Superdex 200 10 300 gl column, by GE Healthcare (1 mentions)
Dawn heleos, by Wyatt Technology (1 mentions)
Astra software, by Wyatt Technology (1 mentions)
B 324, by Büchi (1 mentions)
Sugar pak, by Waters Corporation (1 mentions)
Acquity uplc h class system, by Waters Corporation (1 mentions)
Acetonitrile, by Merck Group (1 mentions)
Acetone, by Tedia (1 mentions)
Vanquish system, by Thermo Fisher Scientific (1 mentions)
Heparin, by Merck Group (1 mentions)
Uv detector, by Waters Corporation (1 mentions)
27 protocols using refractive index detector
1
Characterization of Acrylate-Terminated Polymers
2
Quantification of Ions and Metabolites in MFCs
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For the quantification of Na and Nm, the samples in the anolytes were analyzed using a high-performance liquid chromatography (HPLC) system equipped with a UV detector at 263 nm. The 0.02 M KH2PO4–acetonitrile (90:10, v/v) was used as mobile phase flowing at 1.0 ml min−1 through the Grace Apollo C8 column (4.6 mm × 250 mm, particle size: 5 µm), which was incubated at 28 °C. Lactate in the anolytes was quantified by HPLC with an organic acid column (Aminex HPX-87H Column, 300 mm × 7.8 mm, Bio-Rad) incubated at 65 °C and a refractive index detector (Waters, Corp.). The mobile phase was 0.005 M H2SO4 at a flow rate of 0.6 ml min−1. Samples were centrifuged at 12,000×g for 1 min to remove cells and then filtered with 0.22-μm syringe filters (Nylon) before injection. For the quantification of riboflavin, the samples in the MFCs supernatant were first centrifuged (35,000×g for 20 min) and filtered (0.22 μm), and the eluted medium were detected by a liquid chromatography-tandem mass spectrometer (LC-MS) (Agilent LCMS-1290-6460) in positive ion mode using a Waters XBridge C8 column (2.1 mm × 100 mm; particle size: 3.5 μm).
3
Quantifying Soluble Carbohydrates in Fruits and Leaves
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Soluble carbohydrate of fruits and leaves were measured based on the method used by Niu et al.29 with slight modifications. Triplicate tissues were prepared for fruits and leaves at each stage of development. Frozen samples (about 3 g FW) were ground in liquid nitrogen and extracted in 70% (V/V) ethanol of 6 ml and was incubated in water at 35 °C for 20 min, then it was centrifuged (6,500 g, at 4 °C for 15 min), and the supernatant was transferred to a 25 ml flask, the volume was topped up with 80% ethanol. The ethanolic extract was concentrated by nitrogen gas and the concentrate was reconstituted with 1 ml of ultrapure water. The soluble carbohydrates were separated by HPLC using a CARBOSep CHO-620 CA carbohydrate column (6×250 mm, Shoko Co., Ltd., Tokyo, Japan) dissolved in double-distilled H2O, following the manufacturer's instructions, and a refractive index detector (Waters, Sunnyvale, CA, USA). The column temperature was 80 °C and the flow rate was 0.6 ml min−1 with water as the eluent. Sucrose, glucose, and fructose were determined through their retention times and quantified based on standards.
4
Depolymerization Assay of Extracellular Polysaccharide
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To demonstrate that recombinant the Dep42 had depolymerase activity for the polysaccharide, we performed a depolymerization assay using size exclusion high-performance liquid chromatography (SEC-HPLC). The purified EPS was dissolved in 50 mM Na2HPO4 (pH 7.0) to a final concentration of 0.5 mg/ml and incubated with Dep42 (10 μg/ml) or SUMO (10 μg/ml) at 37°C for 30 min. After incubation, the reactions were stopped by heating at 100°C for 10 min. Subsequently, the mixture was analyzed by an HPLC instrument (Waters, USA) equipped with a TSKgel G5000 PWxl analytical column (7.8 mm ID × 300 mm) (Tosoh Corporation Bioscience, Japan) and a refractive index detector (Waters, USA). The column was run with sodium phosphate buffer at pH 7.0 as the mobile phase at 1 ml/min and 30°C.
5
Characterization of Crosslinked Polymeric Nanoparticles
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1H NMR was used to characterize the polymer structures and the degree of crosslinking. The polymers were dissolved in deuterated DMSO (DMSO-d6) and nanoparticles were first lyophilized and then dissolved in DMSO-d6 for characterization using a Bruker 500 MHz NMR and analyzed using TopSpin 3.6 software. To determine the degree of crosslinking, the acrylate peaks, which are used to form crosslinks upon exposure to UV in the presence of the photoinitiator Irg, were integrated and normalized to protons peaks in the PBAE backbone.
GPC was used to characterize the molecular weight of polymers relative to linear polystyrene standards using a refractive index detector (Waters, Milford, MA). To measure the molecular weight after crosslinking, nanoparticles were formed as described above, and then lyophilized to remove aqueous buffer. Prior to characterization, samples were dissolved in butylated hydroxytoluene-stabilized THF, filtered through a 0.2 μm polytetrafluoroethylene filter.
GPC was used to characterize the molecular weight of polymers relative to linear polystyrene standards using a refractive index detector (Waters, Milford, MA). To measure the molecular weight after crosslinking, nanoparticles were formed as described above, and then lyophilized to remove aqueous buffer. Prior to characterization, samples were dissolved in butylated hydroxytoluene-stabilized THF, filtered through a 0.2 μm polytetrafluoroethylene filter.
6
Polymer Molecular Weight Analysis
7
Quantifying Riboflavin and Lactate
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The supernatant of the MFCs was centrifuged (35000 rpm for 20 min) and filtered (0.22 μm). The eluted samples were analyzed for RF and lactate using liquid chromatograph-tandem mass spectrometer (Agilent LCMS-1290−6460) and high-performance liquid chromatography, respectively. For the quantification of RF, a Waters XBridge C8 column (2.1 mm × 100 mm; particle size: 3.5 μm) was used in positive ion mode. Lactate was quantified by an organic acid column (Aminex HPX-87H Column, 300 mm × 7.8 mm, Bio-Rad) with a refractive index detector (Waters, Corp.) at 65 °C. H2SO4 (5 mM) acted as the mobile phase at a flow rate of 0.6 ml min−1.
8
l-Ribose Fermentation by C. tropicalis
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l -Ribose fermentation with C. tropicalis K1 CoSTP2 LsaAraA AcLRI was performed in a 250-mL Erlenmeyer flask containing 50 mL of fermentation medium at 30 °C with 200 rpm. The fermentation medium was contained 30 g/L or 50 g/L of l -arabinose, 20 g/L of glucose, 10 g/L of yeast extract, 5 g/L of KH2PO4, and 0.2 g/L of MgSO4·7H2O.
The concentrations of glucose,l -arabinose, l -ribulose, and l -ribose were analyzed by high-performance liquid chromatography equipped with a refractive index detector (Waters, Milford, MA, USA). The samples were separated by a Sugar-Pak I column (Waters) with degassed DDW at a flow rate of 0.3 mL/min. Cell growth was determined at 600 nm spectrophotometrically. One A600 was equivalent to 0.474 g (dry cell weight)/L.
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The concentrations of glucose,
9
Quantifying Glucose, Fructose, and Free Amino Nitrogen
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Glucose and fructose were analyzed using high performance liquid chromatography (Waters, UK) by injection of 10 μL of supernatant on an Aminex HPX-87H column (Bio-Rad, USA) and eluted isocratically with 5 mM H2SO4 at 0.5 mL min−1 and 65°C. Detection was performed by a refractive index detector (Waters, UK) at 35°C.
Free amino nitrogen (FAN) concentration was measured based on the ninhydrin reaction method as described by Lie [26 ]. Glycine was used as reference compound.
Free amino nitrogen (FAN) concentration was measured based on the ninhydrin reaction method as described by Lie [26 ]. Glycine was used as reference compound.
10
HPLC Analysis of Metabolite Concentrations
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Extracellular concentrations of glucose, acetic acid, lactic acid, and ethanol were measured by high performance liquid chromatography (HPLC) (Waters, Bedford, MA, USA) using an HPX-87H column (Bio-Rad Aminex, Hercules, CA, USA). The column was eluted with 5 mM H2SO4 at a constant rate of 0.6 mL/min at 50°C. A refractive index detector and a UV dual absorbance detector (Waters, Bedford, MA, USA) were used for detection of the metabolites. UV detection was carried out at 210 nm.
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