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Ultimate 3000 rs system

Manufactured by Thermo Fisher Scientific
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The UltiMate 3000 RS system is a high-performance liquid chromatography (HPLC) system designed for analytical and preparative applications. It offers precise flow and pressure control, advanced temperature regulation, and reliable sample handling. The system is capable of delivering a wide range of flow rates and operating pressures, making it suitable for a variety of HPLC techniques and applications.

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Lab products found in correlation

Zorbax eclipse plus c18 column, by Agilent Technologies (3 mentions) Hypersil gold column, by Thermo Fisher Scientific (2 mentions) Tsq vantage mass spectrometer, by Thermo Fisher Scientific (1 mentions) Accucore c18 column, by Thermo Fisher Scientific (1 mentions) Thermo scientific q exactive orbitrap mass spectrometer, by Thermo Fisher Scientific (1 mentions) Thermo hypersil gold column, by Thermo Fisher Scientific (1 mentions) Sequencing grade trypsin, by Merck Group (1 mentions) Ods 3, by Thermo Fisher Scientific (1 mentions) Hypersil gold c18, by Thermo Fisher Scientific (1 mentions) Xcalibur 3, by Thermo Fisher Scientific (1 mentions) Q exactive mass spectrometer, by Thermo Fisher Scientific (1 mentions) Mzcloud, by Thermo Fisher Scientific (1 mentions) High resolution mass spectrometer, by Thermo Fisher Scientific (1 mentions) Lpg 3400rs pump, by Thermo Fisher Scientific (1 mentions) Dad 3000rs, by Thermo Fisher Scientific (1 mentions) Wps 3000trs, by Thermo Fisher Scientific (1 mentions) Hanna ph meter, by Hanna Instruments (1 mentions) Ltq ion trap mass spectrometer, by Thermo Fisher Scientific (1 mentions) Kinetex c18 column, by Phenomenex (1 mentions) Ltq xl linear ion trap mass spectrometer, by Thermo Fisher Scientific (1 mentions)

23 protocols using ultimate 3000 rs system

1

Anthocyanin and Polyphenol Quantification in Apple Skin

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Anthocyanin content was measured by HPLC. For the more comprehensive analysis of the BBX1 over-expressing lines, HPLC was used to detect anthocyanins and polyphenols. Ground apple skin tissue (100 mg) was freeze-dried for 12 hours. Acidified (0.5% HCL) methanol (5x volume) was added to the tissue and placed in the dark for 3 hours to extract. Samples were centrifuged at 16,000 rpm for 4 minutes, then supernatant transferred to a new tube for vacuum evaporation. Extract was then re-suspended in 500 µL of 20% methanol, filtered into a new tube and vacuum evaporated. Extract was then finally re-suspended in 20 µL methanol for analysis. Extract was analysed on a Dionex Ultimate 3000 RS system as previously described by Dare et al.69 (link).
Plunkett B.J., Henry-Kirk R., Friend A., Diack R., Helbig S., Mouhu K., Tomes S., Dare A.P., Espley R.V., Putterill J, & Allan A.C. (2019). Apple B-box factors regulate light-responsive anthocyanin biosynthesis genes. Scientific Reports, 9, 17762.
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2

Reductive Monitoring of Malonato Complexes

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Reduction of complexes 1b* and 5b* (13C=O labeled 1b and 5b) by sodium ascorbate and by glutathione was monitored by 1H13C HMBC NMR spectroscopy at ambient temperature. Solutions of the compounds (2 mM) were prepared in D2O/H2O (1:9); sodium ascorbate (30 mM) or glutathione (30 mM), respectively, was added, and 1H13C HMBC NMR spectra were recorded for 6 days. The reduction was monitored by following the decrease of intensity of malonato CH2/13C=O cross peaks from the complexes at 4.12/174.8 ppm, 4.06/174.6 ppm and 3.38/175.1 ppm for 1b*, and at 4.20/174.7 ppm, 4.16/174.3 ppm, 4.09/174.9 ppm, and 3.39/175.1 ppm for 5b* relative to the increase of signals of the reduced species at 3.52/178.0 ppm and 3.10/176.8 ppm for 1b* and at 3.59/178.1 ppm and 3.09/176.3 ppm for 5b*.
In addition, compounds 4a and 4b (0.5 mM) were incubated with a 15-fold excess of sodium ascorbate in water and PBS solution (pH 7.4) at 37 °C, and their reduction was followed by means of RP-HPLC over 24 h. The analysis was performed on a Dionex Ultimate 3000 RS system, controlled by the Dionex Chromeleon 6.80 software. The chromatographic conditions were as follows: Poroshell 120 SB C18 column (2.1 mm × 150 mm, 2.7 μm); injection volume, 2 μL; flow rate, 0.2 mL/min; temperature of the column, 25 °C; UV–vis detection set up at 210 nm; mobile phase consisted of 0.1% TFA water/MeOH (95:5).
Varbanov H.P., Göschl S., Heffeter P., Theiner S., Roller A., Jensen F., Jakupec M.A., Berger W., Galanski M, & Keppler B.K. (2014). A Novel Class of Bis- and Tris-Chelate Diam(m)inebis(dicarboxylato)platinum(IV) Complexes as Potential Anticancer Prodrugs. Journal of medicinal chemistry, 57(15), 6751-6764.
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3

UHPLC Analytical Optimization for Compounds

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A separate Dionex UltiMate 3000 RS system was used for analytical optimization without coupling to MS. Chromatographic conditions were the same as for qualitative UHPLC analysis, with the additional use of a Kinetex C18 column, 100 × 2.1 mm, 2.6 μm column and a Kinetex PFP column, 100 × 2.1 mm, 2.6 μm (both Phenomenex, Torrance, CA, USA). Samples of isolated compounds dissolved in methanol (1 mg/mL) were injected at 1 µL; a second dry ethanolic extract in methanol (5 mg/mL) had an injection volume of 2 µL.
Alperth F., Schneebauer A., Kunert O, & Bucar F. (2023). Phytochemical Analysis of Pinus cembra Heartwood—UHPLC-DAD-ESI-MSn with Focus on Flavonoids, Stilbenes, Bibenzyls and Improved HPLC Separation. Plants, 12(19), 3388.
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4

HPLC-MS Analysis of Ethanolic Extracts

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Analyses of the first ethanolic extract, combined fractions and reference substances were conducted on a Dionex UltiMate 3000 RS system equipped with a DAD detector and coupled to an LTQ XL linear ion-trap mass spectrometer with ESI ion source (allThermo Fisher Scientific, Waltham, MA, USA). A Zorbax Eclipse Plus C18 column, 2.1 × 100 mm and 1.8 µm particle size (Agilent, Santa Clara, CA, USA) served as stationary phase, while the mobile phase was made up of water +0.1% formic acid (A) and acetonitrile (B). Gradient elution at a flow rate of 0.25 mL/min started at 20% B, rising to 100% B at 20.0 min, followed by a plateau of 100% B to 23.0 min and a drop back to 20% B at 23.5 min, which was kept stable until 29 min for re-equilibration. Column temperature was 35 °C. Injection volumes were 2 µL for dry ethanolic extract in methanol (5 mg/mL) and 1 µL for reference substances dissolved in methanol (1 mg/mL) as well as subfractions of varying concentrations.
DAD-UV spectra were recorded in a wavelength range of 190 to 700 nm. Mass spectral detection was performed in the range of m/z 50 to 2000 amu, with conditions set as follows: source heater temperature 250 °C, sheath gas flow 50 arb (arbitrary units), auxiliary gas flow 8 arb, source voltage 4.0 kV for ESI negative mode and 4.2 kV for ESI positive mode.
Alperth F., Schneebauer A., Kunert O, & Bucar F. (2023). Phytochemical Analysis of Pinus cembra Heartwood—UHPLC-DAD-ESI-MSn with Focus on Flavonoids, Stilbenes, Bibenzyls and Improved HPLC Separation. Plants, 12(19), 3388.
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5

UHPLC Analytical Optimization for Compounds

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A separate Dionex UltiMate 3000 RS system was used for analytical optimization without coupling to MS. Chromatographic conditions were the same as for qualitative UHPLC analysis, with the additional use of a Kinetex C18 column, 100 × 2.1 mm, 2.6 μm column and a Kinetex PFP column, 100 × 2.1 mm, 2.6 μm (both Phenomenex, Torrance, CA, USA). Samples of isolated compounds dissolved in methanol (1 mg/mL) were injected at 1 µL; a second dry ethanolic extract in methanol (5 mg/mL) had an injection volume of 2 µL.
Alperth F., Schneebauer A., Kunert O, & Bucar F. (2023). Phytochemical Analysis of Pinus cembra Heartwood—UHPLC-DAD-ESI-MSn with Focus on Flavonoids, Stilbenes, Bibenzyls and Improved HPLC Separation. Plants, 12(19), 3388.
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6

HPLC-MS Analysis of Ethanolic Extracts

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Analyses of the first ethanolic extract, combined fractions and reference substances were conducted on a Dionex UltiMate 3000 RS system equipped with a DAD detector and coupled to an LTQ XL linear ion-trap mass spectrometer with ESI ion source (allThermo Fisher Scientific, Waltham, MA, USA). A Zorbax Eclipse Plus C18 column, 2.1 × 100 mm and 1.8 µm particle size (Agilent, Santa Clara, CA, USA) served as stationary phase, while the mobile phase was made up of water +0.1% formic acid (A) and acetonitrile (B). Gradient elution at a flow rate of 0.25 mL/min started at 20% B, rising to 100% B at 20.0 min, followed by a plateau of 100% B to 23.0 min and a drop back to 20% B at 23.5 min, which was kept stable until 29 min for re-equilibration. Column temperature was 35 °C. Injection volumes were 2 µL for dry ethanolic extract in methanol (5 mg/mL) and 1 µL for reference substances dissolved in methanol (1 mg/mL) as well as subfractions of varying concentrations.
DAD-UV spectra were recorded in a wavelength range of 190 to 700 nm. Mass spectral detection was performed in the range of m/z 50 to 2000 amu, with conditions set as follows: source heater temperature 250 °C, sheath gas flow 50 arb (arbitrary units), auxiliary gas flow 8 arb, source voltage 4.0 kV for ESI negative mode and 4.2 kV for ESI positive mode.
Alperth F., Schneebauer A., Kunert O, & Bucar F. (2023). Phytochemical Analysis of Pinus cembra Heartwood—UHPLC-DAD-ESI-MSn with Focus on Flavonoids, Stilbenes, Bibenzyls and Improved HPLC Separation. Plants, 12(19), 3388.
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7

Liquid Chromatography-Mass Spectrometry Protocol

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Liquid chromatography was performed with an UltiMate™ 3000 RS system, coupled to a TSQ Vantage mass spectrometer (Thermo Fisher Scientific, USA). Separation was performed using a Syncronise C18 column of 50 mm length and 4.6 mm internal diameter, with a particle size of 1.7 μm (Thermo Fisher Scientific, USA).
Gajardo G., López-Muñoz R., Plaza A., Uberti B., Sarmiento J., Morán G, & Henríquez C. (2019). Tamoxifen in horses: pharmacokinetics and safety study. Irish Veterinary Journal, 72, 5.
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8

Quantification of Allithiamine using LC-MS

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Prior to MS a reversed-phase chromatographic separation was used. 10 μl of standard solution of allithiamine was injected to a Thermo Scientific Ultimate 3000 RS system equipped with a Thermo Accucore C18 column (100/2.1, 2.6 μm). Eluent A (500 ml of water containing 10 ml of methanol, 0.5 ml of formic acid and 2.5 mM of ammonium formate (pH 2.7) and eluent B (500 ml of methanol containing 10 ml of water, 0.5 ml of formic acid and 2.5 mM of ammonium formate) were mixed in 50–50%. Flow rate was 0.2 ml/min. The chromatography system was coupled to a Thermo Scientific Q Exactive Orbitrap mass spectrometer (Thermo Fisher Scientific Inc., Waltham, USA) equipped with an electrospray ionization source (ESI). The mass spectrometer was operated with the following parameters: capillary temperature 320 °C, spray voltage 4.0 kV, the resolution was set to 70000. The mass range scanned was 100–1000 m/z. The maximum injection time was 100 ms. Sheath gas flow rate was 32 arb, aux gas flow rate was 7 arb.
Fragmentation of allithiamine was studied in positive ionisation mode at 40 Normalized Collision Energy (NCE).
Biro A., Gál F., Hegedűs C., Batta G., Cziáky Z., Peitl B., Stündl L., Gyémánt G, & Remenyik J. (2018). Isolation of allithiamine from Hungarian red sweet pepper seed (Capsicum annuum L.). Heliyon, 4(12), e00997.
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9

HPLC-MS Analysis of RR-L Compound

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200 mg RR-L extraction was added with 1 ml methanol, whirled for 10 min, centrifuged at 4°C for 10 min, filtered with a 0.22 μm filter membrane, and subjected to Ultimate 3000 RS system (Thermo Fisher Scientific, MA, USA) equipped with a Thermo Hypersil GOLD column (φ 2.1 × 100 mm, 1.9 μm). The MS spectra were acquired by a Q Executive high-resolution mass spectrometer (Thermo Fisher Scientific, MA, USA). The mobile phases were (A) 0.1% formic acid in water (B) and 0.1% formic acid in acetonitrile, and the gradient elution program was as follows (time/B%): 0–1 min, 2%; 1–5 min, 2%–20%; 5–10 min, 20–50%; 10–15 min, 50–80%; 15–20 min, 80–95%; 20–25 min, 95%; 26–30 min, 2%. The chromatographic analysis was performed at 35°C with a flow rate of 0.3 mL/min and an injection volume of 15 μL. The mass spectrometer parameters were as follows: spray voltage was set at 3.8 kV at positive mode. The capillary temperature was set at 300°C. Argon was used as the collision gas. Nitrogen was used as sheath gas and aux gas. Aux gas heater temperature was set to be 350°C. The chromatogram was analyzed by CD 2.1 software.
Ling X., Wang X., Li J., Zhang T., Zhou X., Zhou Y, & Kang J. (2021). Shu-Di-Huang and Gan-Cao Herb Pair Restored the Differentiation Potentials of Mesenchymal Stem Progenitors in Treating Osteoporosis via Downregulation of NF-κB Signaling Pathway. Evidence-based Complementary and Alternative Medicine : eCAM, 2021, 7795527.
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10

Proteomics Analysis of Desulfovibrio Oralis

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Samples from the same three replicate D. oralis cultures used for the proinflammatory assay were also used for proteomic analysis, conducted as detailed in reference 100 (link). Cell pellets were resuspended in 4% sodium deoxycholate (SDC) in 100 mM ammonium bicarbonate (ABC) and ultrasonically disrupted. The crude protein extract was clarified by centrifugation and reduced with 10 mM dithiothreitol (DTT), and cysteine residues were blocked with 30 mM iodoacetamide (IAA). The proteins were then collected on top of a 10-kDa cutoff spin column filter. D. oralis spent culture medium samples (~12 ml) were concentrated on a 5-kDa filter. Collected proteins were washed with ABC and digested with sequencing-grade trypsin (Sigma). Peptides were collected by centrifugation, acidified to 1% formic acid (FA) followed by extraction with ethyl acetate, and concentrated. The peptide mixture was analyzed by an automated, two-dimensional liquid chromatography-tandem mass spectrometry (2D-LC-MS/MS) system (100 (link)) using the Ultimate 3000 RS system in-line with a Q Exactive Plus (QE+) mass spectrometer (Thermo, Fisher Scientific).
Cross K.L., Chirania P., Xiong W., Beall C.J., Elkins J.G., Giannone R.J., Griffen A.L., Guss A.M., Hettich R.L., Joshi S.S., Mokrzan E.M., Martin R.K., Zhulin I.B., Leys E.J, & Podar M. (2018). Insights into the Evolution of Host Association through the Isolation and Characterization of a Novel Human Periodontal Pathobiont, Desulfobulbus oralis. mBio, 9(2), e02061-17.
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