The analysis of samples was performed using HPLC instrument (Agilent) equipped with a Rezex RPM-Monosaccharide Pb2+ column (300 × 7.8 mm; particle size 8 μm) at 80 °C. Millipore water was used as an eluent with a flow rate of 0.6 mL/min. The refractive index detector (RID) with a cell temperature of 40 °C was used to detect the products. Before analysis, the samples were filtered through 0.22 μm syringe filter, which was then injected for the analysis. The calibration curve was plotted with the standard compounds for calculating the yield of products. The Gas chromatography (GC) equipped with HP-5 column was used for the analysis of the organic layer of the reaction mixture. The Flame Ionization Detector with the temperature 280 °C was used for the examination of verity of products.
Hplc instrument
Manufactured by Agilent Technologies
Sourced in United States
The HPLC (High-Performance Liquid Chromatography) instrument is a laboratory equipment used for the separation, identification, and quantification of various chemical compounds in a mixture. It consists of a solvent delivery system, an injection port, a separation column, and a detector. The instrument operates by pumping a liquid mobile phase through the column, which contains a stationary phase, and the analytes in the sample are separated based on their interactions with the stationary phase.
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Lab products found in correlation
Eclipse xdb c18, by Agilent Technologies (1 mentions)
Atlantis dc18 column, by Waters Corporation (1 mentions)
Avance 3 500 mhz, by Bruker (1 mentions)
Zetasizer nano zs instrument, by Malvern Panalytical (1 mentions)
Dsc q100, by TA Instruments (1 mentions)
Cytation 3 imaging reader, by Agilent Technologies (1 mentions)
Jem 1400 tem, by JEOL (1 mentions)
Ivis lumina xr imaging system, by PerkinElmer (1 mentions)
Avance 400 mhz spectrometer, by Bruker (1 mentions)
F 2500 fluorescence spectrophotometer, by Hitachi (1 mentions)
Qtrap 5500 mass spectrometer, by AB Sciex (1 mentions)
C18 reversed phase column, by Thermo Fisher Scientific (1 mentions)
Acetonitrile, by Merck Group (1 mentions)
2489 hplc system, by Waters Corporation (1 mentions)
Mcp 200 automatic polarimeter, by Anton Paar (1 mentions)
Accurate mass q tof lc ms 6520 instrument, by Agilent Technologies (1 mentions)
Sephadex lh 20, by Cytiva (1 mentions)
Avance spectrometer, by Bruker (1 mentions)
Fdu 1200, by Eyela (1 mentions)
Diode array detector, by Agilent Technologies (1 mentions)
18 protocols using hplc instrument
1
Analysis of Monosaccharides via HPLC-RID
2
Quantification of Bioactive Compounds Using HPLC
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Appropriate volumes of the drugs were obtained and immersed in two volumes of water for 30 min. Then, 10 volumes of water were added, boiled and then filtered. Eight volumes of water were added to the residues, boiled and filtered. The liquid obtained by both the filtration were combined and concentrated to a concentration of 1 g/ml crude drug. Exactly 5 ml of the extraction was added to an evaporating dish, evaporated dry, and then 50% methanol was added to obtain a final volume of 10 ml. The liquid was mixed then filtered, and the filtered liquid was further filtered through a 0.45-µm microfiltration membrane to obtain the solution for the experiment. Gradient elution was performed with the mobile phase of acetonitrile (A) and 0.1% phosphoric acid solution (B). The flow velocity used was 1.0 ml/min, the column temperature was 40°C, the sample size was 20 µl and the determine wavelengths were as follows: 0–11 min, 290 nm; 11–27 min, 320 nm; 27–45 min, 230 nm; and 45–80 min, 270 nm. An HPLC instrument (Agilent Technologies) was used for measuring the concentration of the liquid (Figs. 1 and 2 ).
3
HPLC Quantification of Resveratrol and Polydatin
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HPLC detection was conducted using an Agilent HPLC instrument coupled with Eclipse XDB-C18 (4.6 × 150 mm 5 μm) at 306 nm at 30 °C. HPLC grade acetonitrile and water containing 0.1% formic acid were used as mobile phase A and B. The gradient elution condition was shown in Table 7 . Standard curves were prepared using concentration (C) and peak area (A) of resveratrol and polydatin standard compound, respectively. The transformation rate was calculated using the equation [25 (link)]: polydatin: A = 42895C − 234.38 (R² = 0.9999, linear range: 0.010–0.800 mg/mL); resveratrol: A = 56488C + 152.94, (R² = 0.9998, linear range: 0.005–0.400 mg/mL); emodin: A = 54984C + 227.71, R² = 0.9999, (linear range: 0.001 mg/mL–0.400 mg/mL).
Dried powder of different plant tissues was weighed accurately to about 50 mg in an Eppendorf tube and 1.5 mL methanol was added. The mixture was sonicated for 30 min and followed by centrifugation at 12,000 rpm for 10 min. The supernatant was withdrawn and filtered using a 0.45 μM filter; 10 μL was used for HPLC analysis.
Transformation rate (%) = (increased resveratrol concentration/original polydatin concentration) × (390.40/228.25) × 100, in which 228.25 and 390.40 are the molecular weight of resveratrol and polydatin, respectively.
Dried powder of different plant tissues was weighed accurately to about 50 mg in an Eppendorf tube and 1.5 mL methanol was added. The mixture was sonicated for 30 min and followed by centrifugation at 12,000 rpm for 10 min. The supernatant was withdrawn and filtered using a 0.45 μM filter; 10 μL was used for HPLC analysis.
Transformation rate (%) = (increased resveratrol concentration/original polydatin concentration) × (390.40/228.25) × 100, in which 228.25 and 390.40 are the molecular weight of resveratrol and polydatin, respectively.
4
HPLC Analysis of Caffeine in Tea
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For the chromatographic analysis, an HPLC instrument (Agilent Technologies, Tokyo, Japan, 1260) equipped with a C18 column (reverse phase, 3.9 × 150 mm, 10 µm 125 Å (Waters; Nova-Pak, Milford, MA, United State of America) was used. The injection volume was 10 μL and the column temperature was 38 °C with a 15 min running time (stop time) for the caffeine standards and tea samples. The signal of caffeine was monitored at a wavelength of 273 nm using a diode array detector (DAD) signal. The caffeine stock standard (analytical grade, Sigma-Aldrich, Darmstadt, Hesse, Germany) solutions were prepared by dissolving 0.1 g of sample in 100 mL of distilled water. The working standard solutions of caffeine were prepared by suitable dilution from the stock solution, and the stock solution was then stored in the refrigerator for further analysis. Separation of the caffeine was achieved with water–acetonitrile (85:15 v/v) with 0.1% formic acid as the mobile phase at a flow rate of 1 mL min−1. All the chemicals, reagents and solvents used were of analytical grade.
5
HPLC Analysis of Bioactive Compounds in DJS Extract
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The contents albiflorin, paeoniflorin, z-ligustilide, decursin, and nodakenin in the DJS water extract were analyzed using an HPLC instrument (Agilent Technologies, USA) with a Atlantis dC18 column (4.6 × 250 mm, 5 μm; Waters, USA). The mobile phase consisted of the solvents, distilled water (A) and acetonitrile with 0.1% formic acid (B). The following gradient was used: 0 min, A : B 90 : 10 (v/v); 20 min, A : B 75 : 25; 25 min, A : B 75 : 25; 30 min, A : B 50 : 50; 45 min, A : B 20 : 80; and 60 min, A : B 0 : 100. The mobile phase flow rate was 1.0 mL/min, the column temperature was 30°C, the injection volume was 10 μL, and UV detection was at 230 and 330 nm.
6
HPLC Analysis of Phenolic Compounds in FRO
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FRO was analyzed using an HPLC instrument (Agilent Technologies, Santa Clara, CA, USA) with Hichrome HPLC columns (5 μm, 250 mm × 4.6 mm; Hichrome, Theale, UK). The flow rate used was 0.3 mL/min, and the injection volume used was 10 μL. Additionally, 100% methanol was used as the solvent, and a detection wavelength of 254 nm was set. Polyphenol standards (gallic acid, fisetin, quercetin, and kaempferol) were used to characterize the phenolic compounds in the FRO extract. The mobile phase consisted of 0.1% formic acid in water (solvent A) and 100% methanol (solvent B) and was delivered at a flow rate of 0.7 mL/min. The following gradient conditions were used: 0–17 min, 100% B; 17–20 min, 100% B; 20–23 min, 0% B; 23–30 min, 0% B. The detection wavelength used was 254 nm, and the injection volume for all polyphenol samples was 10 μL.
7
Comprehensive Characterization of Polymers
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A Bruker AVANCE III (500 MHz) nuclear magnetic resonance (NMR) instrument was used to collect 1H and 13C spectra using CDCl3 as the solvent. Size exclusion chromatography (SEC) measurements were obtained using a Shimadzu HPLC instrument equipped with an Agilent column connected to the Shimadzu refractive index detector with N, N-dimethylformamide (DMF) as eluent, and poly(methyl methacrylate) (PMMA) standard calibration. Differential scanning calorimetry (DSC) was performed on a TA Instruments Q100 DSC under nitrogen at 20 mL/min. A temperature-controlled Cary5000 UV-vis spectrometer was used for the turbidimetric assay of the synthesized polymers. A 0.5°C min-1 heating rate was applied, and a quartz cuvette with a path length of 1.0 cm was used. Fluorescence spectroscopy was performed using a Perkin-Elmer LS 50 BL luminescence spectrometer. The size and distribution of the particles were measured through dynamic light scattering (DLS) using the Malvern Zetasizer Nano ZS instrument equipped with a He-Ne laser (633 nm) and a 173° backscatter detector. Transmission electron microscopy (TEM) analysis was conducted using a JEM-1400+ TEM (JEOL United States Inc., MA) with 2% phosphotungstic acid stain. A Biotek Cytation 3 imaging reader was used to perform cell viability and cellular uptake measurements.
8
NQO1-Targeted NIR Probe Evaluation
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All chemicals and solvents used in syntheses were purchased from Sigma- Aldrich or Fisher Scientific and used without further purification. NQO1 was purchased from Sigma-Aldrich (D1515). NQO1 siRNA (h) (sc-37139) and NQO1 (A180) (sc-32793) antibodies were purchased from Santa Cruz. All cell lines were obtained from ATCC (American type cell culture collection) The 1H, 13C NMR spectra were recorded on a Bruker-Avance 400 MHz Spectrometer. Chemical shifts (δ) are reported in ppm. ESI mass spectra were recorded on AB sciex QTRAP 5500 mass spectrometer. High-performance liquid chromatography (HPLC) was performed on an Agilent HPLC instrument. Peaks in NMR spectra are listed as singlet (s), doublet (d), triplet (t), or multiplet (m), and coupling constants (J) are reported in hertz (Hz). Fluorescence spectra were recorded on a Hitachi F-2500 Fluorescence spectrophotometer in a 10 mm standard cell with both excitation and emission slit widths of 10 nm. The incubation of NQO1 with NIR-ASM in the presence of NADH was carried out on a shaker at 37 °C. IVIS Lumina XR Imaging system (Caliper Life Sciences, Inc.) was used for the in vivo imaging. The BD LSRFortessa™ cell analyzer was used for the flow cytometric analysis. A Nikon multiphoton microscope equipped with NIS-Elements C acquisition and analysis software was used for the fluorescence microscopy.
9
HPLC Quantification of Fungal Metabolites
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For HPLC assays, the leaves and fungal endophytes were dried at 110°C for 10 min, then at 60°C for 2 or 3 days to a constant weight. The leaves and fungal endophytes were ground into a powder, and 100 mg of the dried powder was accurately weighed, extracted with 1 mL of methanol (contains 0.1% of hydrochloric acid) for 12 h, and sonicated for 60 min. The extracts were centrifuged for 10 min at 4,000 rpm at 4°C, and supernatants were filtered with 0.45-μm filter columns. The filtrates (10 μL) were uploaded for analyses on a reversed-phase C18 column (Thermo) using an HPLC instrument (Agilent, USA). The elution phase was acetonitrile (Sigma, St. Louis, MO, USA): methanol: water (A: B: C) = 95:0.5:4.5 and detected with a UV detector at 254 nm. The elution speed was 1 mL/min, and the column temperature was 30°C. Samples were eluted using the gradient procedures illustrated in S1 Table . The content of metabolites (mg/g) was quantified using catechin as a standard with an R2 = 0.999.
10
Grape Callus Metabolite Extraction
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Ten milligram of freezing dried grape callus powder were accurately weighed and extracted with 500 μL of 60% methanol (contains 0.1% of hydrochloric acid) for 2 hours in an ultrasonic cleaner. Extracts were centrifuged briefly at 10000 rpm and supernatants were then filtered with 0.45 um filter columns. Ten micro liter of extracts were loaded and metabolites in grape cells were separated by reverse C18 column on a HPLC instrument (Agilent, USA) with 30 °C of column temperature. Elution phase is acetonitrile: water: formic acid = 35: 65: 0.1, with the elution speed of 1 mL/min, and detected with a UV detector at 254, 263 and 280 nm, respectively. Samples were eluted with the gradient procedures as illustrated in supplementary table (S1 Table ).
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